Neonates and nursing infants are special with regard to immune development and vulnerability to infectious diseases. Although breast-feeding is essential to modulate and prime immune defenses, vaccines (an interventional prophylaxis) are crucial to prevent and control infectious diseases. During nursing, the type of feeding influences infants' natural defenses (including gut colonization) and their response to vaccines, both through cell-mediated immunity and specific antibody production. Given the variety and combination of vaccine components (antigens and excipients, preservative thimerosal, and aluminum adjuvants) and route of administration, there is a need to examine the role of infant feeding practices in intended and nonintended outcomes of vaccination. Maternal factors related to milk constituents (nutrients and pollutants) and feeding practices can affect response to vaccines. Collectively, studies that compared type of feeding (or used breast-feeding-adjusted statistical models) showed significant influence on some vaccines taken during infancy. Nurslings deprived of the full benefit of breast-feeding could have altered immune responses affecting vaccine outcome. In the absence of studies elucidating neurodevelopment (including excitoxicity) and immunotoxicity issues, vaccination practices should promote and support breast-feeding.

Neurodevelopment as Gesell development scores (GDSs) in relation to mercury exposure in infants (<6 months of age) of one urban center and two rural villages, respectively, of fisherman and cassiterite miners. Mean total hair-Hg (HHg) concentrations of infants from Itapuã (3.95 ± 1.8 ppm) were statistically (P = 0.0001) different from those of infants from Porto Velho (3.84 ± 5.5 ppm) and Bom Futuro (1.85 ± 0.9 ppm). Differences in vaccine coverage among these populations resulted in significantly higher (P = 0.0001) mean ethylmercury (EtHg) exposure in urban infants (150 μg) than in infants from either village (41.67 μg, Itapuã; 42.39 μg, Bom Futuro). There was an inverse significant (Spearman r =-0.2300; P = 0.0376) correlation between HHg and GDS for infants from Porto Velho, but not for the rural infants from Bom Futuro (Spearman r = 0.1336; P = 0.0862) and Itapuã (Spearman r = 0.1666; P = 0.5182). Logistic regression applied to variables above or below the median GDS showed that EtHg exposure (estimated probability =-0.0157; P = 0.0070) and breastfeeding score (estimated probability =-0.0066; P = 0.0536) score were significantly associated with GDS. Conclusion. In nurslings whose mothers are exposed to different levels of fish-MeHg (HHg), a higher score of neurological development at six months was negatively associated with exposure to additional TCV-EtHg. Results should be interpreted with caution because of unaccounted variables.

Metformin (dimethyl-biguanide) is an insulin-sensitizing agent that lowers fasting plasma-insulin concentration, wherefore it's wide use for patients with a variety of insulin-resistant and prediabetic states, including impaired glucose tolerance. During pregnancy it is a further resource for reducing first-trimester pregnancy loss in women with the polycystic ovary syndrome. We tested metformin genotoxicity in cells of Chinese hamster ovary, CHO-K1 (chromosome aberrations; comet assays) and in mice (micronucleus assays). Concentrations of 114.4 μg/mL and 572 μg/mL were used in in vitro tests, and 95.4 mg/kg, 190.8 mg/kg and 333.9 mg/kg in assaying. Although the in vitro tests revealed no chromosome aberrations in metaphase cells, DNA damage was detected by comet assaying after 24 h of incubation at both concentrations. The frequency of DNA damage was higher at concentrations of 114.4 μg/mL. Furthermore, although mortality was not observed in in vitro tests, the highest dose of metformin suppressed bone marrow cells. However, no statistically significant differences were noted in micronuclei frequencies between treatments. In vitro results indicate that chronic metformin exposure may be potentially genotoxic. Thus, pregnant woman undergoing treatment with metformin should be properly evaluated beforehand, as regards vulnerability to DNA damage.

Birth weight data (22,012 live-births) from a public hospital in Porto Velho (Amazon) was used in multiple statistical models to assess the effects of forest-fire smoke on human reproductive outcome. Mean birth weights for girls (3,139 g) and boys (3,393 g) were considered statistically different (p-value < 2.2e-16). Among all models analyzed, the means were considered statistically different only when treated as a function of month and year (p-value = 0.0989, girls and 0.0079, boys). The R (2) statistics indicate that the regression models considered are able to explain 65 %(girls) and 54 %(boys) of the variation of the mean birth weight.

β-glucan is an important polysaccharide due to its medicinal properties of stimulating the immune system and preventing chronic diseases such as cancer. The aim of the present study was to determine the anticlastogenic effect of β-glucan in cells exposed to ultraviolet radiation (UV). Chromosome aberration assay was performed in drug-metabolizing cells (HTC) and non drug-metabolizing cells (CHO-K1 and repair-deficient CHO-xrs5), using different treatment protocols. Continuous treatment (UV + β-glucan) was not effective in reducing the DNA damage only in CHO-xrs5 cells. However, the pre-treatment protocol (β-glucan before UV exposition) was effective in reducing DNA damage only in CHO-K1 cells. In post-treatment (β-glucan after UV exposition) did not show significative anticlastogenic effects, although there was a tendency toward prevention. The data suggest that β-glucan has more than one action mechanism, being capable of exerting desmutagenic as well as bio-antimutagenic action. The findings also suggest that the presence of the xenobiotic metabolizing system can reduce the chemopreventive capacity of β-glucan. Therefore, these results indicate that β-glucan from Saccharomyces cerevisiae can be used in the prevention and/or reduction of DNA damage.

It is thought that high linear energy transfer (LET) radiation induces more complex DNA damage than low LET particles, specifically clustered DNA damage that causes cells to repair DNA double strand breaks (DSB) more slowly and leads to severe biological consequences. The present study aimed to investigate the role of exogenously added glutathione (GSH) on (12)C-beam (287 keV/mum) and (7)Li-beam (60 keV/mum) induced chromosome aberration (CA) formation, particularly on exchange aberration formation. In order to characterize the role of GSH in the joining of DNA DSBs, we induced DNA lesions with bleomycin (Blem) in conjunction with either high- or low-LET radiation (X-rays) since the chemistry of the free DNA ends created by Blem and X-rays is similar. CHO cells were exposed to reduced GSH at a concentration of 2mM for 3h before radiation. Treatment with Blem (20mug/ml) was carried out for 2h before the cells were exposed to radiation. Our results show that the frequency of chromosomal aberration increases with increased LET. Heavy ion exposed cells show a higher frequency of CA over time than do X-irradiated cells. An analysis of the first post-irradiation mitosis of exposed CHO cells shows that high-LET radiation induces more breaks than exchange-type aberrations and exogenous GSH has no influence on high-LET radiation induced DNA damage. The DNA lesions induced by low LET radiation interact relatively strongly with Blem-induced lesions whereas interaction between Blem and high LET radiations was poor. This could be attributed to differences in repair kinetics and qualitative differences in the DNA lesions induced by Blem and high-LET radiation.

Beta-sitosterol is an important phytosterol found in plant food. It has been shown to have antiproliferative effects on cancers of the colon, breast, and prostate, but its effect on stomach cancer cells in vitro is unknown. Proliferation, cytotoxicity, and apoptosis in SGC-7901 human stomach cancer cells were examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, clone formation, lactate dehydrogenase (LDH) leakage assay, acridine orange (AO)/ethidium bromide (EB) double staining, 4',6-diamidine-2'-phenylindole dihydrochloride (DAPI) staining, comet assay, and Western blotting. The results showed that beta-sitosterol suppresses the proliferation and induces the cell cytotoxicity of SGC-7901 stomach cancer cells in a time- and dose-dependent manner. Cells treated with different concentrations of beta-sitosterol also showed changes typical of apoptosis: morphological changes, DNA damage, increased expression of pro-caspase-3 and bax (p < 0.05), and activation of pro-caspase-3 and suppression of bcl-2 expression (p < 0.05). This study therefore revealed that beta-sitosterol significantly inhibits the growth and induces the apoptosis of SGC-7901 human stomach cancer cells in vitro. The decrease of the bcl-2/bax ratio and DNA damage may be the critical mechanisms of apoptosis induced by beta-sitosterol in SGC-7901 human stomach cancer cells.

BACKGROUND Piper sarmentosum, locally known as kaduk is belonging to the family of Piperaceae. It is our interest to evaluate their effect on human hepatoma cell line (HepG2) for the potential of anticarcinogenic activity. RESULTS The anticarcinogenic activity of an ethanolic extract from Piper sarmentosum in HepG2 and non-malignant Chang's liver cell lines has been previously determined using (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide)(MTT) assays, where the IC50 value was used as a parameter for cytotoxicity. The ethanolic extract that showed anticarcinogenic properties in HepG2 cells had an IC50 of 12.5 mug mL-1, while IC50 values in the non-malignant Chang's liver cell line were greater than 30 mug mL-1. Apoptotic morphological changes in HepG2 cells were observed using an inverted microscope and showed chromatin condensation, cell shrinkage and apoptotic bodies following May-Grunwald-Giemsa's staining. The percentage of apoptotic cells in the overall population (apoptotic index) showed a continuously significant increase (p < 0.05) in 12.5 mug mL-1 ethanolic extract-treated cells at 24, 48 and 72 hours compared to controls (untreated cells). Following acridine orange and ethidium bromide staining, treatment with 10, 12 and 14 mug mL-1 of ethanolic extracts caused typical apoptotic morphological changes in HepG2 cells. Molecular analysis of DNA fragmentation was used to examine intrinsic apoptosis induced by the ethanolic extracts. These results showed a typical intrinsic apoptotic characterisation, which included fragmentation of nuclear DNA in ethanolic extract-treated HepG2 cells. However, the non-malignant Chang's liver cell line produced no DNA fragmentation. In addition, the DNA genome was similarly intact for both the untreated non-malignant Chang's liver and HepG2 cell lines. CONCLUSION Therefore, our results suggest that the ethanolic extract from P. sarmentosum induced anticarcinogenic activity through an intrinsic apoptosis pathway in HepG2 cells in vitro.

J Oral Pathol Med (2008)Background: Bracken fern (Pteridium aquilinum) has been consumed by humans and animals for centuries. However, its consumption is associated with a high incidence of cancer in the upper digestory tract of different species. Although the oral cavity is the first site of contact with ingested toxic substances, the interaction of bracken fern composites with oral cell lines has not yet been studied. Methods: In order to study the biological responses of oral cells exposed to bracken fern, we evaluated the genotoxic and cytotoxic effects of a bracken fern aqueous extract in oral cell lines. Human submandibular gland (HSG) and human oral epithelium cells (OSCC-3) cells were treated with three different concentrations of the extract. DNA damage was determined by the comet assay, and cellular morphology was examined by light microscopy. Apoptotic changes were evaluated by transmission electron microscopy and TUNEL assay. Results: The comet assay revealed that the extract was genotoxic for both cell lines but the results were not dose-dependent. The morphological and ultrastructural analyses showed that the extract caused conspicuous alterations in both cell types: uncommon chromatin condensation, nuclear picnosis, cellular volume decrease, nuclear envelope disruption, formation of numerous vacuoles of different sizes and apoptotic bodies. The TUNEL assay confirmed apoptosis induction. Conclusions: These results demonstrate that the extract was cytotoxic to HSG and OSCC-3 cells, and that cellular degeneration occurred mainly by apoptosis. We believe that oral cells could trigger apoptosis after bracken fern induced DNA damage, in order to avoid the malignant transformation.

We examined the genotoxic and mutagenic effects of a crude extract of Clusia alata (a potential medicinal plant) on peripheral leukocyte and bone marrow cells of mice, using the comet and chromosome aberration assays. Extracts at doses of 1000, 1500 and 2000 mg/kg were administered by gavage, and a positive control, N-nitroso-N-ethylurea (50 mg/kg) was injected intraperitoneally. Peripheral blood leukocytes were collected 4 and 24 h after the treatments for the comet assay, and bone marrow cells were collected 24 h after the treatments, for the chromosome aberration assay. The comet assay showed that C. alata extract causes an increase in damage to DNA in the peripheral blood leukocytes, but it was significant only with the 2000 mg/kg dose after 24 h; the extract also induced a small but significant increase in the mean number of chromosome aberrations in the bone marrow cells at doses of 1500 and 2000 mg/kg. No evidence of a significant decrease in the mitotic index was observed. Acute consumption of high concentrations of C. alata extract produced some mutagenic effects in bone marrow cells.

BACKGROUND Lumiracoxib is a highly selective cyclooxygenase-2 (COX-2) inhibitor with antiinflammatory, analgesic and antipyretic activities comparable with class specific drugs, but with much improved gastrointestinal safety. No studies have examined lumiracoxib for antitumorigenic activity on human nonsmall cell lung cancer cell lines in vitro or its possible molecular mechanisms. METHODS The antiproliferative effect of lumiracoxib alone or combined with docetaxol on A549 and NCI-H460 lines was assessed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Drug-drug interactions were analyzed using the coefficient of drug interaction (CDI) to characterize the interactions as synergism, additivity or antagonism. Morphological changes were observed by acridine orange fluorescent staining. Extent of apoptosis was determined by flow cytometry. RESULTS Lumiracoxib (15 - 240 micromol/L) has an inhibitory effect on the proliferation of A549 and NCI-H460 cell lines in concentration- and time-dependent manners with the IC50 values of 2597 micromol/L and 833 micromol/L, respectively. The synergistic effect was prominent when lumiracoxib (15 - 240 micromol/L) was combined with docetaxol (0.2 - 2 micromol/L)(CDI < 1). Fluorescent staining showed that lumiracoxib could induce apoptosis in A549 and NCI-H460 cells. Lumiracoxib treatment also caused an increase of the sub-G1 fraction in each cell line and resulted in an increase of G0/G1-phase cells and a decrease of S-phase cells. CONCLUSIONS Lumiracoxib had antiproliferative effect on the human nonsmall cell lung cancer cell lines A549 and NCI-H460 and had a significant synergy with docetaxol, which may be related to apoptotic induction and cell cycle arrest.

The genotoxic effects of tryptophol (indole-3-ethanol), an aromatic alcohol and known secondary metabolite of the opportunistic yeast Candida albicans and other Candida spp., were studied using a battery of short-term assays on human white blood cells in vitro. The concentration range of tryptophol tested was 0.25 mM to 2.00 mM. Lymphocyte viability and induction of apoptosis/necrosis were studied by simultaneous use of a fluorescent assay with ethidium bromide and acridine orange. Levels of primary DNA damage and dynamics of DNA repair were evaluated using the alkaline comet assay while the levels and nature of residual DNA damage were assessed by the analysis of structural chromosome aberrations, the sister chromatid exchange test and the cytokinesis-block micronucleus assay. The results obtained suggest cytotoxic, cytostatic and genotoxic effects of the tryptophol treatment in vitro that were mainly dose-dependent. The type and the extent of DNA lesions detected in tryptophol-treated samples indicate the possibility that observed damage is mediated by highly reactive aldehyde metabolite and/or free radicals produced by treatment. The results show that mortality of lymphocytes in tryptophol-treated samples was primarily caused by apoptosis. The generation of additional DNA strand breaks and cytogenetic consequences (chromosome aberrations, sister chromatid exchanges and micronuclei), as observed in this study, sustain the possibility that tryptophol toxicity is mediated by the formation of DNA cross-links and aldehyde-protein adducts. In conclusion, this preliminary study elucidates only a part of tryptophol toxicity to human cells. Because current evidence is not sufficient to obtain information relevant for human risk assessment, further in vitro and in vivo studies are essential in order to clarify remaining issues, especially to elucidate the exact mechanisms and nature of the damage produced following treatment as well to estimate possible interindividual variability in genotoxic responses to the chemical.

Cylindrospermopsin (CYN) is a cyanobacterial alkaloid that has been implicated in outbreaks of human morbidity and animal mortality. The principal mode of action for CYN is inhibition of protein and glutathione synthesis, and its toxicity seems to be mediated by cytochrome P-450-generated metabolites. It was also shown that CYN might be responsible for tumor initiation in animals; nevertheless, mechanisms leading to CYN-induced carcinogenesis are scarce and equivocal. The aim of the present study was to investigate the impact of metabolic activation on CYN-induced DNA damage. The effect of different doses of CYN (0.05-2mug/ml) on DNA damage was determined in CHO-K1 cells after 3, 16 and 21h of the treatment. The chromosome aberration assay with and without metabolic activation was applied to evaluate the clastogenic activity of CYN and its metabolite(s). In addition, the occurrence of apoptosis and necrosis was estimated by the annexin method using flow cytometry. The results revealed that CYN is not clastogenic in CHO-K1 cells irrespective of S9 fraction-induced metabolic activation. However, CYN significantly decreases the frequencies of mitotic indices and decreases proliferation irrespective of metabolic activation system. CYN increases the frequency of necrotic cells in a dose- and time-dependent manner, whereas it has a very slight impact on apoptosis. Moreover, the presence of metabolic activation influences a susceptibility to necrotic cell death but not an apoptotic one.

Telomeres and telomerase appear to participate in the repair of broken DNA ends produced by oxidative damage. Arsenite is an environmental contaminant and a potent human carcinogen, which induces oxidative stress on cells via the generation of reactive oxygen species affecting cell viability and chromosome stability. It promotes telomere attrition and reduces cell survival by apoptosis. In this study, we used mouse embryonic fibroblasts (MEFs) from mice lacking telomerase RNA component (mTERC(-/-) mice) with long (early passage or EP) and short (late passage or LP) telomeres to investigate the extent of oxidative damage by comparing the differences in DNA damage, chromosome instability, and cell survival at 24 and 48 h of exposure to sodium arsenite (As3+; NaAsO2). There was significantly high level of DNA damage in mTERC(-/-) cells with short telomeres as determined by alkaline comet assay. Consistent with elevated DNA damage, increased micronuclei (MN) induction reflecting gross genomic instability was also observed. Fluorescence in situ hybridization (FISH) analysis revealed that increasing doses of arsenite augmented the chromosome aberrations, which contributes to genomic instability leading to possibly apoptotic cell death and cell cycle arrest. Microarray analysis has revealed that As3+ treatment altered the expression of 456 genes of which 20% of them have known functions in cell cycle and DNA damage signaling and response, cell growth, and/or maintenance. Results from our studies imply that short dysfunctional telomeres impair the repair of oxidative damage caused by arsenite. The results will have implications in risk estimation as well as cancer chemotherapy.