Mobile phone specific radiation disturbs cytokinesis and causes cell death but not acute chromosomal damage in buccal cells: Results of a controlled human intervention study.

Several human studies indicate that mobile phone specific electromagnetic fields may cause cancer in humans but the underlying molecular mechanisms are currently not known. Studies concerning chromosomal damage (which is causally related to cancer induction) are controversial and those addressing this issue in mobile phone users are based on the use of questionnaires to assess the exposure. We realized the first human intervention trial in which chromosomal damage and acute toxic effects were studied under controlled conditions. The participants were exposed via headsets at one randomly assigned side of the head to low and high doses of a UMTS signal (n = 20, to 0.1 W/kg and n = 21 to 1.6 W/kg Specific Absorption Rate) for 2 h on 5 consecutive days. Before and three weeks after the exposure, buccal cells were collected from both cheeks and micronuclei (MN, which are formed as a consequence of structural and numerical chromosomal aberrations) and other nuclear anomalies reflecting mitotic disturbance and acute cytotoxic effects were scored. We found no evidence for induction of MN and of nuclear buds which are caused by gene amplifications, but a significant increase of binucleated cells which are formed as a consequence of disturbed cell divisions, and of karyolitic cells, which are indicative for cell death. No such effects were seen in cells from the less exposed side. Our findings indicate that mobile phone specific high frequency electromagnetic fields do not cause acute chromosomal damage in oral mucosa cells under the present experimental conditions. However, we found clear evidence for disturbance of the cell cycle and cytotoxicity. These effects may play a causal role in the induction of adverse long term health effects in humans.

Impact of mobile phone-specific electromagnetic fields on DNA damage caused by occupationally relevant exposures: results of ex vivo experiments with peripheral blood mononuclear cells from different demographic groups.

The aim of this study was to investigate if age and body mass of humans have an impact on the DNA-damaging properties of high-frequency mobile phone-specific electromagnetic fields (HF-EMF, 1950 MHz, universal mobile telecommunications system, UMTS signal) and if this form of radiation has an impact on the genotoxic effects of occupationally relevant exposures. Pooled peripheral blood mononuclear cells (PBMC) from three groups [young normal weight, young obese (YO), and older age normal weight individuals] were exposed to different doses of HF-EMF (0.25, 0.5, and 1.0 W/kg specific absorption rate-SAR) and simultaneously or sequentially to different chemicals which cause DNA damage (CrO3, NiCl2, benzo[a]pyrene diol epoxide-BPDE, and 4-nitroquinoline 1-oxide-4NQO) via different molecular mechanisms. We found no difference in regard to the background values in the three groups but a significant increase of DNA damage (81% without and 36% with serum) in cells from old participants after radiation with 1.0 W/kg SAR 16 h. In combined treatment experiments we found no impact of the UMTS signal on chemically induced DNA damage in the different groups in general. However, a moderate decrease of DNA damage was seen in simultaneous treatment experiments with BPDE and 1.0 W/kg SAR in the YO group (decline 18%). Taken together our findings indicate that HF-EMF cause DNA damage in PBMC from older subjects (69.1 years). Furthermore, they show that the radiation does not increase induction of DNA damage by occupationally relevant chemicals.

Gallic acid, a common dietary phenolic protects against high fat diet induced DNA damage.

PURPOSE: Aim of the study was to find out if gallic acid (GA), a common phenolic in plant foods, prevents obesity induced DNA damage which plays a key role in the induction of overweight associated cancer. METHODS: Male and female C57BL6/J mice were fed with a low fat or a high fat diet (HFD). The HFD group received different doses GA (0, 2.6-20 mg/kg b.w./day) in the drinking water for 1 week. Subsequently, alterations of the genetic stability in blood and inner organs were monitored in single cell gel electrophoresis assays. To elucidate the underlying molecular mechanisms: oxidized DNA bases, alterations of the redox status, lipid and glucose metabolism, cytokine levels and hepatic NF-κB activity were monitored. RESULTS: HFD fed animals had higher body weights; increased DNA damage and oxidation of DNA bases damage were detected in colon, liver and brain but not in blood and white adipose tissue. Furthermore, elevated concentrations of insulin, glucose, triglycerides, MCP-1, TNF-α and NF-κB activity were observed in this group. Small amounts of GA, in the range of human consumption, caused DNA protection and reduced oxidation of DNA bases, as well as biochemical and inflammatory parameters. CONCLUSIONS: Obese animals have increased DNA damage due to oxidation of DNA bases. This effect is probably caused by increased levels of glucose and insulin. The effects of GA can be explained by its hypoglycaemic properties and indicate that the consumption of GA-rich foods prevents adverse health effects in obese individuals.

Low doses of widely consumed cannabinoids (cannabidiol and cannabidivarin) cause DNA damage and chromosomal aberrations in human-derived cells.

Cannabidiol (CBD) and cannabidivarin (CBDV) are natural cannabinoids which are consumed in increasing amounts worldwide in cannabis extracts, as they prevent epilepsy, anxiety, and seizures. It was claimed that they may be useful in cancer therapy and have anti-inflammatory properties. Adverse long-term effects of these drugs (induction of cancer and infertility) which are related to damage of the genetic material have not been investigated. Therefore, we studied their DNA-damaging properties in human-derived cell lines under conditions which reflect the exposure of consumers. Both compounds induced DNA damage in single cell gel electrophoresis (SCGE) experiments in a human liver cell line (HepG2) and in buccal-derived cells (TR146) at low levels (≥ 0.2 µM). Results of micronucleus (MN) cytome assays showed that the damage leads to formation of MNi which reflect chromosomal aberrations and leads to nuclear buds and bridges which are a consequence of gene amplifications and dicentric chromosomes. Additional experiments indicate that these effects are caused by oxidative base damage and that liver enzymes (S9) increase the genotoxic activity of both compounds. Our findings show that low concentrations of CBD and CBDV cause damage of the genetic material in human-derived cells. Furthermore, earlier studies showed that they cause chromosomal aberrations and MN in bone marrow of mice. Fixation of damage of the DNA in the form of chromosomal damage is generally considered to be essential in the multistep process of malignancy, therefore the currently available data are indicative for potential carcinogenic properties of the cannabinoids.

Use of HuH6 and other human-derived hepatoma lines for the detection of genotoxins: a new hope for laboratory animals?

Cell lines which are currently used in genotoxicity tests lack enzymes which activate/detoxify mutagens. Therefore, rodent-derived liver preparations are used which reflect their metabolism in humans only partly; as a consequence misleading results are often obtained. Previous findings suggest that certain liver cell lines express phase I/II enzymes and detect promutagens without activation; however, their use is hampered by different shortcomings. The aim of this study was the identification of a suitable cell line. The sensitivity of twelve hepatic cell lines was investigated in single cell gel electrophoresis assays. Furthermore, characteristics of these lines were studied which are relevant for their use in genotoxicity assays (mitotic activity, p53 status, chromosome number, and stability). Three lines (HuH6, HCC1.2, and HepG2) detected representatives of five classes of promutagens, namely, IQ and PhIP (HAAs), B(a)P (PAH), NDMA (nitrosamine), and AFB1 (aflatoxin), and were sensitive towards reactive oxygen species (ROS). In contrast, the commercially available line HepaRG, postulated to be a surrogate for hepatocytes and an ideal tool for mutagenicity tests, did not detect IQ and was relatively insensitive towards ROS. All other lines failed to detect two or more compounds. HCC1.2 cells have a high and unstable chromosome number and mutated p53, these features distract from its use in routine screening. HepG2 was frequently employed in earlier studies, but pronounced inter-laboratory variations were observed. HuH6 was never used in genotoxicity experiments and is highly promising, it has a stable karyotype and we demonstrated that the results of genotoxicity experiments are reproducible.

Genotoxic and Cytotoxic Effects in Exfoliated Buccal and Nasal Cells of Chromium and Cobalt Exposed Electroplaters.

Results of a number of studies indicate that electroplaters have increased cancer risks as a consequence of exposure to genotoxic metals such as chromium (VI) and nickel. These effects may be due to induction of damage of the genetic material which plays a key role in the etiology of cancer, and it was found that workers in galvanization factories exhibited increased levels of DNA damage. The aim of the present study was to investigate genetic stability in workers of a bright plating factory who are exposed to chromium (Cr) and cobalt (Co). Exfoliated cells were collected from the buccal and nasal mucosa of workers (n = 42) and matched controls (n = 43) and analyzed for induction of micronuclei (MN) which are formed as a consequence of chromosomal aberrations. In addition, other nuclear anomalies namely nuclear buds (Nbuds) which are formed as a consequence of gene amplification and markers indicating different stages of cell death (condensed chromatin, karyorrhexis, karyolysis, and pyknosis) were also assessed. No evidence was noted for induction of MN, but significantly increased rates of Nbuds in cells from both, buccal and nasal mucosa, were found. Parameters which are indicative for cytotoxic effects were more pronounced in nasal cells and rose with duration of employment period. Overall, our findings indicated that no apparent chromosomal damage occurred in bright electroplaters. However, data demonstrated that acute cytotoxic effects may lead to inflammations and/or lesions in epithelia of the respiratory tract of the workers.

Genotoxic properties of XLR-11, a widely consumed synthetic cannabinoid, and of the benzoyl indole RCS-4.

Aim of this study was the investigation of the genotoxic properties of XLR-11 [1-(5-fluoropentyl)-1H-indol-3-yl](2,2,3,3-tetramethylcyclopropyl)methanone, a widely consumed synthetic cannabinoid (SC), and of the benzoyl indole RCS-4 (4-methoxyphenyl)(1-pentyl-1H-indol-3-yl)methanone). We characterized the DNA-damaging properties of these drugs in different experimental systems. No evidence for induction of gene mutations was detected in bacterial (Salmonella/microsome) tests, but clear dose-dependent effects were found in in vitro single cell gel electrophoresis (SCGE) assays with human lymphocytes and with buccal- and lung-derived human cell lines (TR-146 and A-549). These experiments are based on the determination of DNA migration in an electric field and enable the detection of single- and double-strand breaks and apurinic sites. Furthermore, we found that both drugs induce micronuclei which are formed as a consequence of chromosomal aberrations. The lack of effects in SCGE experiments with lesion-specific enzymes (FPG, Endo III) shows that the DNA damage is not caused by formation of oxidatively damaged bases; experiments with liver enzyme homogenates and bovine serum albumin indicate that the drugs are not converted enzymatically to DNA-reactive intermediates. Furthermore, results with buccal- and lung-derived human cells show that gaseous treatment of the cells under conditions which reflect the exposure situation in drug users may cause damage of the genetic material in epithelia of the respiratory tract. Since DNA instability is involved in the etiology of cancer, these findings can be taken as an indication that consumption of the SCs may cause tumors in the respiratory tract of consumers.

Cytotoxic and genotoxic activities of waters and sediments from highway and parking lot runoffs.

The genotoxicity of water and sediment samples from stormwater treatment systems and water from urban highway runoff was tested in the Salmonella/microsome assays with Salmonella typhimurium, micronucleus assay (Trad-MN) with plants and with human-derived liver cells (HepG2), or comet assay with HepG2. Cytotoxicity of water samples was studied using either reactive oxygen species (ROS) generation, cell proliferation or dye exclusion assay in HepG2. Concentrations of several contaminants in the tested samples were also measured. Results suggested that urban highway runoff exposed to severe vehicle traffic emissions caused genotoxic effects in comet assay and in Trad-MN assays. Sediments induced either mutagenic effects in strain YG1024 or genotoxic effects in Trad-MN assay. These effects could be due to the presence of nitro-polycyclic aromatic hydrocarbons (NPAHs) which possess carcinogenic and mutagenic properties. Influent and effluents of stormwater treatment systems did not induce genotoxic activity or effects on HepG2 cell viability; however, the influents were able to induce ROS generation and cell proliferation in HepG2 cells. As the methods require a sterile filtration of the water samples, this could have also removed particulate-associated polycyclic aromatic hydrocarbons (PAHs) and resulted in a less pronounced induction of genotoxicity, as would be expected by PAH contamination.

Investigation of the in vitro toxicological properties of the synthetic cannabimimetic drug CP-47,497-C8.

Cannabicyclohexanol (CP-47,497-C8) is a representative of a group of cannabimimetic cyclohexylphenols which is added to herbal mixtures as a cannabis substitute since 2008. Although in the beginning CP-47,497-C8 was the main ingredient of "Spice" and similar products, it was partly replaced by aminoalkylindole-type cannabinoid receptor agonists like JWH-018, JWH-073 or JWH-250, but never completely disappeared from the market. Since information on its toxicological properties is scarce, we investigated the effects of the drug in human derived cell lines. The cytotoxic effects were studied in a panel of assays (SRB, XTT, LDHe and NR tests) in a buccal derived (TR146) and a liver derived (HepG2) cell line. The strongest effects were seen in the two former assays at levels ≥ 7.5 µM indicating that the compound interferes with protein synthesis and causes membrane damage. In additional comet assays, DNA damage was detected at levels ≥10 µM. Experiments with lesion specific enzymes showed that these effects are not due to oxidative damage of DNA bases. The negative findings obtained in Salmonella/microsome assays and the positive results of micronucleus tests with the cell lines indicate that the compound does not cause gene mutations but acts on the chromosomal level. In contrast to other synthetic cannabinoids, no indication for estrogenic/antiestrogenic properties was seen in a luciferase assay with bone marrow derived U2-OS cells. In conclusion, our findings show that the drug has only weak cytotoxic properties. However, the induction of chromosomal damage indicates that it may cause adverse effects in users due to its impact on the stability of the genetic material.

Acute toxic and genotoxic activities of widely used cytostatic drugs in higher plants: Possible impact on the environment.

Cytostatic drugs are highly toxic pharmaceuticals and it was repeatedly postulated that they may cause adverse effects in ecosystems. The acute toxic and genotoxic properties of these drugs have not been adequately investigated in higher plants so far; therefore, we studied the most widely used drugs (5-flurouracil, 5FU; etoposide, Et; cisplatin, CisPt; carboplatin, CaPt; vincristine sulfate, VinS and cyclophosphamide monohydrate, CP) in micronucleus (MN) assays with meiotic pollen tetrad cells of Tradescantia and with root cells from Allium cepa. MNi are formed as a consequence of chromosome breaks and aneuploidy. We monitored also the acute toxic properties of the drugs, i.e. inhibition of cell division (mitotic indices and retardation of root growth) in the latter species. All compounds caused in both indicator plants genotoxic effects. The order of genotoxic potencies expressed as NOELs in µM was CisPt (0.1)≥ Et (0.5)>CP (1.0)>CaPt (10)>5FU (30)>VinS (100) in Tradescantia. A similar order was seen in Allium MN but Et was less active (5.0µM). Four compounds caused alterations of the mitotic indices under the present conditions namely CisPt (0.5), Et (10.0), 5FU (10.0) and VinS (100). Inhibition of root growth decreased in the order CisPt (0.5)>Et (1.0)≥VinS (1.0)>5FU (5.0)>CaPt (33.0)>CP (>1000). Comparisons of the NOELs with the predicted environmental concentrations (PEC) show that the latter values are at least 5 orders of magnitude lower and indicate that it is unlikely that their release in the environment may cause adverse effects in higher plants. However, it is notable that the levels of both platinum compounds and of 5FU in hospital effluents may reach levels which may induce damage of the genetic material.

Tetraploidy as a metastable state towards malignant cell transformation within a systemic approach of cancer development.

Tetraploidy, a condition in which a cell has four homologous sets of chromosomes, may be a natural physiological condition or pathophysiological such as in cancer cells or stress induced tetraploidisation. Its contribution to cancer development is well known. However, among the many models proposed to explain the causes, mechanisms and steps of malignant cell transformation, only few integrate tetraploidization into a systemic multistep approach of carcinogenesis. Therefore, we will i) describe the molecular and cellular characteristics of tetraploidy; ii) assess the contribution of stress-induced tetraploidy in cancer development; iii) situate tetraploidy as a metastable state leading to cancer development in a systemic cell-centered approach; iiii) consider knowledge gaps and future perspectives. The available data shows that stress-induced tetraploidisation/polyploidisation leads to p53 stabilisation, cell cycle arrest, followed by cellular senescence or apoptosis, suppressing the proliferation of tetraploid cells. However, if tetraploid cells escape the G1-tetraploidy checkpoint, it may lead to uncontrolled proliferation of tetraploid cells, micronuclei induction, aneuploidy and deploidisation. In addition, tetraploidization favors 3D-chromatin changes and epigenetic effects. The combined effects of genetic and epigenetic changes allow the expression of oncogenic gene expression and cancer progression. Moreover, since micronuclei are inducing inflammation, which in turn may induce additional tetraploidization, tetraploidy-derived genetic instability leads to a carcinogenic vicious cycle. The concept that polyploid cells are metastable intermediates between diploidy and aneuploidy is not new. Metastability denotes an intermediate energetic state within a dynamic system other than the system's state at least energy. Considering in parallel the genetic/epigenetic changes and the probable entropy levels induced by stress-induced tetraploidisation provides a new systemic approach to describe cancer development.

Impact of high (1950 MHz) and extremely low (50 Hz) frequency electromagnetic fields on DNA damage caused by occupationally relevant exposures in human derived cell lines.

Epidemiological studies indicate that electromagnetic fields (EMF) are associated with cancer in humans. Exposure to mobile phone specific high frequency fields (HF-EMF) may lead to increased glioma risks, while low frequency radiation (LF-EMF) is associated with childhood leukemia. We studied the impact of HF-EMF (1950 MHz, UMTS signal) on DNA stability in an astrocytoma cell line (1321N1), and the effect of LF-EMF (50 Hz) in human derived lymphoma (Jurkat) cells. To find out if these fields affect chemically induced DNA damage, co-exposure experiments were performed. The cells were exposed to HF-EMF or LF-EMF and treated simultaneously and sequentially with mutagens. The compounds cause DNA damage via different molecular mechanisms, i.e. pyrimidine dimers which are characteristic for UV light (4-nitroquinoline 1-oxide, 4NQO), bulky base adducts (benzo[a]pyrene diolepoxide, BPDE), DNA-DNA and DNA-protein cross links and oxidative damage (NiCl2, CrO3). DNA damage was measured in single cell gel electrophoresis (comet) assays. We found a moderate reduction of basal and 4NQO-induced DNA damage in the astrocytoma line, but no significant alterations of chemically induced DNA migration by the HF and LF fields under all other experimental series. The biological consequences of the moderate reduction remain unclear, but our findings indicate that acute mobile phone and power line specific EMF exposures do not enhance genotoxic effects caused by occupationally relevant chemical exposures.

Use of micronucleus cytome assays with buccal cells for the detection of genotoxic effects: A systematic review and meta-analysis of occupational exposures to metals.

Micronucleus (MN) assays with buccal cells are at present widely used to investigate occupational exposures to genotoxic carcinogens. This article describes their use for the monitoring of metal exposed workers. We found in total 73 relevant articles, in the majority (97 %) increased MN and/or other nuclear anomalies were reported. Most studies were realized in South East Asia and South America. A variety of different occupations was studied including welders, electroplaters, painters, workers in battery recycling and production, tannery workers, dental technicians, miners, workers in foundries and smelters, and also subjects working in waste recycling, glass, aluminum and steel production. In many investigations the effects increased with the duration of the working period. The quality of individual studies was evaluated with a quality score tool. The number of cells was in most studies sufficient and DNA-specific stains were used. However, many studies have shortcomings, e.g. they focused solely on MN formation and did not evaluate anomalies, which provide additional information about the stability of the genetic material and acute cytotoxic effects. Only 35 % of the investigations contain quantitative information about exposures to metals and other toxicants. In 6 of these studies, correlations were observed between the concentrations of specific metals (As, Pb, Cr, Cd) in body fluids and MN frequencies. Taken together, the available data indicate that the MN assay can be used to detect chromosomal damage in metal exposed groups; furthermore, it enables also comparisons between subgroups differing in regard to their exposure and allows an estimation of the efficiency of protective measures. The exposure of workers to metals is currently controlled with chemical analytical measurements only, MN assays with buccal cells could contribute to further improve the safety at workplaces as they reflect the biological consequences including synergistic and antagonistic interactions between toxicants.

Induction of nuclear anomalies in exfoliated buccal cells of coca chewers: results of a field study.

The leaves of coca (Erythroxylum coca var. coca), a South American shrub which contains cocaine, other alkaloids and phenolics are widely used by indigenous populations of the Andes. It is currently not known if coca consumption causes genotoxic effects in humans. This information is important to predict potential long-term toxic effects such as cancer induction. Therefore, the buccal cytome assay was used to analyze oral cells from 45 uni- and bilateral chewers and 23 controls living in the Altiplano of the Peruvian Andes. In total, 123,471 cells were evaluated from chewers and 57,916 from controls. Information concerning the consumption levels and habits and also use of lime were collected with questionnaires. Chewing of the leaves did not induce nuclear anomalies reflecting genetic damage such as micronuclei (MNi) and nuclear buds; in the highest exposure group (but not in the overall group) even a significant decrease in the frequencies of cells with MNi (by 64 %) was observed. However, we found significantly elevated levels of other nuclear anomalies (karyorrhexis and karyolysis) which reflect cytotoxic effects in the coca users. The frequencies of these anomalies increased with the daily consumption and when lime was used to improve the release of the alkaloids. In contrast to other chewing habits (betel, tobacco and khat), consumption of coca leaves does not induce genetic instability in cells from the oral cavity and our findings indicate that no adverse health effects take place in chewers which are associated with DNA damage. However, the significant increase in certain anomalies shows that acute toxic effects are caused by coca consumption.

Impact of Bariatric Surgery on the Stability of the Genetic Material, Oxidation, and Repair of DNA and Telomere Lengths.

Obesity causes genetic instability, which plays a key-role in the etiology of cancer and aging. We investigated the impact of bariatric surgery (BS) on DNA repair, oxidative DNA damage, telomere lengths, alterations of antioxidant enzymes and, selected proteins which reflect inflammation. The study was realized with BS patients (n = 35). DNA damage, base oxidation, BER, and NER were measured before and 1 month and 6 months after surgery with the single-cell gel electrophoresis technique. SOD and GPx were quantified spectrophotometrically, malondealdehyde (MDA) was quantified by HPLC. Telomere lengths were determined with qPCR, and plasma proteome profiling was performed with high-resolution mass spectrophotometry. Six months after the operations, reduction of body weight by 27.5% was observed. DNA damage decreased after this period, this effect was paralleled by reduced formation of oxidized DNA bases, a decline in the MDA levels and of BER and NER, and an increase in the telomere lengths. The activities of antioxidant enzymes were not altered. Clear downregulation of certain proteins (CRP, SAA1) which reflect inflammation and cancer risks was observed. Our findings show that BS causes reduced oxidative damage of DNA bases, possibly as a consequence of reduction of inflammation and lipid peroxidation, and indicate that the surgery has beneficial long-term health effects.

Use of the single cell gel electrophoresis assay for the detection of DNA-protective dietary factors: Results of human intervention studies.

The single cell gel electrophoresis technique is based on the measurement of DNA migration in an electric field and enables to investigate via determination of DNA-damage the impact of foods and their constituents on the genetic stability. DNA-damage leads to adverse effects including cancer, neurodegenerative disorders and infertility. In the last 25 years approximately 90 human intervention trials have been published in which DNA-damage, formation of oxidized bases, alterations of the sensitivity towards reactive oxygen species and chemicals and of repair functions were investigated with this technique. In approximately 50% of the studies protective effects were observed. Pronounced protection was found with certain plant foods (spinach, kiwi fruits, onions), coffee, green tea, honey and olive oil. Also diets with increased contents of vegetables caused positive effects. Small amounts of certain phenolics (gallic acid, xanthohumol) prevented oxidative damage of DNA; with antioxidant vitamins and cholecalciferol protective effects were only detected after intake of doses that exceed the recommended daily uptake values. The evaluation of the quality of the studies showed that many have methodological shortcomings (lack of controls, no calibration of repair enzymes, inadequate control of the compliance and statistical analyses) which should be avoided in future investigations.

Entinostat Enhances the Efficacy of Chemotherapy in Small Cell Lung Cancer Through S-phase Arrest and Decreased Base Excision Repair.

PURPOSE: Acquired chemoresistance is a frequent event in small cell lung cancer (SCLC), one of the deadliest human malignancies. Histone deacetylase inhibitors (HDACi) have been shown to synergize with different chemotherapeutic agents including cisplatin. Accordingly, we aimed to investigate the dual targeting of HDAC inhibition and chemotherapy in SCLC. EXPERIMENTAL DESIGN: The efficacy of HDACi and chemotherapy in SCLC was investigated both in vitro and in vivo. Synergistic drug interactions were calculated based on the HSA model (Combenefit software). Results from the proteomic analysis were confirmed via ICP-MS, cell-cycle analysis, and comet assays. RESULTS: Single entinostat- or chemotherapy significantly reduced cell viability in human neuroendocrine SCLC cells. The combination of entinostat with either cisplatin, carboplatin, irinotecan, epirubicin, or etoposide led to strong synergy in a subset of resistant SCLC cells. Combination treatment with entinostat and cisplatin significantly decreased tumor growth in vivo. Proteomic analysis comparing the groups of SCLC cell lines with synergistic and additive response patterns indicated alterations in cell-cycle regulation and DNA damage repair. Cell-cycle analysis revealed that cells exhibiting synergistic drug responses displayed a shift from G1 to S-phase compared with cells showing additive features upon dual treatment. Comet assays demonstrated more DNA damage and decreased base excision repair in SCLC cells more responsive to combination therapy. CONCLUSIONS: In this study, we decipher the molecular processes behind synergistic interactions between chemotherapy and HDAC inhibition. Moreover, we report novel mechanisms to overcome drug resistance in SCLC, which may be relevant to increasing therapeutic success.

Cytotoxic and DNA-damaging properties of glyphosate and Roundup in human-derived buccal epithelial cells.

Glyphosate (G) is the largest selling herbicide worldwide; the most common formulations (Roundup, R) contain polyoxyethyleneamine as main surfactant. Recent findings indicate that G exposure may cause DNA damage and cancer in humans. Aim of this investigation was to study the cytotoxic and genotoxic properties of G and R (UltraMax) in a buccal epithelial cell line (TR146), as workers are exposed via inhalation to the herbicide. R induced acute cytotoxic effects at concentrations > 40 mg/l after 20 min, which were due to membrane damage and impairment of mitochondrial functions. With G, increased release of extracellular lactate dehydrogenase indicative for membrane damage was observed at doses > 80 mg/l. Both G and R induced DNA migration in single-cell gel electrophoresis assays at doses > 20 mg/l. Furthermore, an increase of nuclear aberrations that reflect DNA damage was observed. The frequencies of micronuclei and nuclear buds were elevated after 20-min exposure to 10-20 mg/l, while nucleoplasmatic bridges were only enhanced by R at the highest dose (20 mg/l). R was under all conditions more active than its active principle (G). Comparisons with results of earlier studies with lymphocytes and cells from internal organs indicate that epithelial cells are more susceptible to the cytotoxic and DNA-damaging properties of the herbicide and its formulation. Since we found genotoxic effects after short exposure to concentrations that correspond to a 450-fold dilution of spraying used in agriculture, our findings indicate that inhalation may cause DNA damage in exposed individuals.

Use of nasal cells in micronucleus assays and other genotoxicity studies.

Genotoxicity experiments with exfoliated nasal mucosa cells are a promising minimally invasive approach for the detection of DNA-damaging compounds in ambient air. Results of single cell gel electrophoresis (SCGE) assays with individual cells and organ cultures from bioptic material show that DNA damage caused by compounds such as nitrosamines, polycyclic aromatic hydrocarbons and pesticides can be detected. Biochemical studies indicate that enzymes involved in the metabolism of environmental mutagens are represented in nasal cells. Several protocols for experiments with nasal cells have been developed and it was shown that formaldehyde, metals, styrene and crystalline silica induce DNA damage in SCGE and/or in micronucleus studies; furthermore, it was also found that polluted urban air causes DNA instability in nasal epithelial cells. Comparisons of these data with results obtained in lymphocytes and buccal cells indicate that nasal cells are in general equally sensitive. Broad variations in the baseline levels, differences of results obtained in various studies as well as the lack of information concerning the impact of confounding factors on the outcome of experiments with these cells indicate the need for further standardisation of the experimental protocols.

Impact of spinach consumption on DNA stability in peripheral lymphocytes and on biochemical blood parameters: results of a human intervention trial.

INTRODUCTION: A controlled intervention trial was conducted to assess the impact of spinach consumption on DNA stability in lymphocytes and on health-related biochemical parameters. METHODS: The participants (n = 8) consumed homogenised spinach (225 g/day/person) over a period of 16 days. DNA migration was monitored in single cell gel electrophoresis-comet assays under standard conditions, which reflect single- and double-strand breaks, after treatment of nuclei with lesion-specific enzymes (formamidopyrimidine glycosylase, FPG and endonuclease III, ENDO III) and after treatment of intact cells with H(2)O(2) before, during and after intervention. RESULTS: While no reduction in DNA damage was observed under standard conditions after different time intervals of spinach intake, other endpoints, namely ROS sensitivity and DNA migration attributable to the formation of oxidatively damaged DNA bases (i.e. pyrimidines-ENDO III-sensitive sites and purines-FPG sensitive sites) were reduced 6 h after consumption of the first portion and after 11 days of continuous consumption. In the case of ENDO III-sensitive sites, also after 16 days, a decrease in comet formation was observed. At the end of a 40 days washout period, the DNA stability parameters were not significantly different from the background values. Other biochemical parameters which were significantly altered by spinach intake were the folate (+27%) and homocysteine (-16%) concentrations in blood, and it was found in an earlier human study that folate may prevent oxidative damage to DNA bases. CONCLUSIONS: Taken together, our results show that moderate consumption of spinach causes protection against oxidative DNA damage in humans and that this phenomenon is paralleled by alterations of health-related biochemical parameters.