Association of exposure to Bisphenol A with obesity and cardiometabolic risk factors in children and adolescents.

In this study, the association of exposure to Bisphenol A (BPA) with obesity and cardiometabolic risk factors was investigated on 132 children and adolescents aged 6-18 years living in Isfahan, Iran. Potential contributors to BPA exposure were assessed by a questionnaire. Total BPA was detected in urine samples of all participants without significant difference in boys and girls. The mean body mass index (BMI) and waist circumference (WC) increased significantly across the BPA tertiles (p for trend = < 0.001). Similar trend was documented for systolic blood pressure (SBP) and diastolic blood pressure (DBP) as well as fasting blood sugar. The risk of obesity was 12.48 times higher in participants in the third tertile of BPA than in others (95% CI: 3.36-46.39, p < 0.001). The current study showed significant association between BPA exposure with obesity and some cardiometabolic risk factors in children and adolescents, however, further longitudinal studies are necessary to evaluate the clinical effects of this finding. Abbreviations: BMI: Body Mass Index; BPA: Bisphenol A; BSTFA: N, O-Bistrifluoroacetamide; CDC: Centers for Disease Control and Prevention; CI: Circumference Interval; DBP: Diastolic Blood Pressure; DLLME: Dispersive liquid-liquid microextraction method; FBS: Fasting Blood Glucose; HDL: high-density lipoprotein cholesterol were; LDL: low-density lipoprotein cholesterol; OR: Odd Ratio; PA: Physical Activity; SBP: Systolic Blood Pressure; TC: total cholesterol; TG: triglycerides; WC: Waist Circumference.

Is there any association between phthalate exposure and precocious puberty in girls?

Considerable increase in the prevalence of precocious puberty (PP) during the last decade has raised a lot of concerns. Some environmental endocrine disruptor chemicals (EDCs), such as phthalate esters, have intrinsic estrogen activity or increase endogenous sex hormone levels leading to PP. This study was conducted to investigate the association between exposure to phthalate esters and PP in a sample of girls. Plasma levels of seven phthalate ester metabolites were measured in 87 girls with PP and 63 age- and sex-matched controls by dispersive liquid-liquid microextraction and GC/MS analysis. History of exposure to main sources of phthalates was obtained by a checklist. Diethyl hexyl phthalate (DEHP) metabolite levels were significantly higher in those with PP than that in controls (p < 0.05), but this difference was not significant for other phthalate metabolites. 30.1% girls with PP and 12.2% of controls had played for more than 2 h/day with plastic toys in their childhood. 65.1% girls with PP and 32.8% of controls have regularly used some cosmetic products. Consumption of bottled water and beverages by those with PP was about twofold higher than that in the control group. A positive correlation was found between bottled ware consumption and plasma concentrations of four phthalate metabolites. The frequency of seafood consumption was not significantly different between the groups studied. Our findings confirm positive association between phthalate exposure and incidence of PP in girls. Control and reduction of children exposure to phthalate esters should be considered as a health priority.

Development of a simple and valid method for the trace determination of phthalate esters in human plasma using dispersive liquid-liquid microextraction coupled with gas chromatography-mass spectrometry.

A new method was developed for the trace determination of phthalic acid esters in plasma using dispersive liquid-liquid microextraction and gas chromatography with mass spectrometry analysis. Plasma proteins were efficiently precipitated by trichloroacetic acid and then a mixture of chlorobenzene (as extraction solvent) and acetonitrile (as dispersive solvent) rapidly injected to clear supernatant using a syringe. After centrifuging, chlorobenzene sedimented at the bottom of the test tube. 1 µL of this sedimented phase was injected into the gas chromatograph for phthalic acid esters analysis. Different factors affecting the extraction performance, such as the type of extraction and dispersive solvent, their volume, extraction time, and the effects of salt addition were investigated and optimized. Under the optimum conditions, the enrichment factors and extraction recoveries were satisfactory and ranged between 820-1020 and 91-97%, respectively. The linear range was wide (50-1000 ng/mL) and limit of detection was very low (1.5-2.5 ng/mL for all analytes). The relative standard deviations for analysis of 1 µg/mL of the analytes were between 3.2-6.1%. Salt addition showed no significant effect on extraction recovery. Finally, the proposed method was successfully utilized for the extraction and determination of the phthalic acid esters in human plasma samples and satisfactory results were obtained.

Association of atmospheric concentrations of polycyclic aromatic hydrocarbons with their urinary metabolites in children and adolescents.

This study aims to determine the atmospheric concentrations of particulate matter 2.5 (PM2.5)-bounded polycyclic aromatic hydrocarbons (PAHs) and their association with their urinary metabolites in children and adolescents. This study was conducted from October 2014 to March 2016 in Isfahan, Iran. We measured 16 species of PAHs bounded to PM2.5 by gas chromatography mass spectrometry (GC/MS) from 7 parts of the city. Moreover, PAH urinary metabolites were measured in 186 children and adolescents, randomly selected from households. Urinary metabolites consisted of 1-hydroxy naphthalene (1-naphthol), 2-hydroxy naphthalene (2-naphthol), 9-hydroxy phenanthrene (9-phenanthrol), and 1-hydroxy pyrene using GC/MS. Considering the short half-lives of PAHs, we measured the metabolites twice with 4 to 6 months of time interval. We found that the ambient concentrations of PAHs were significantly associated with their urinary metabolites. 1-hydroxy naphthalene and 2-hydroxy naphthalene concentrations showed an increase of 1.049 (95% CI: 1.030, 1.069) and 1.047 (95% CI: 1.025, 1.066) for each unit increase (1 ng/m3) in ambient naphthalene. Similarly, 1-hydroxy pyrene showed an increase of 1.009 (95% CI: 1.006-1.011) for each unit increase (1 ng/m3) in ambient pyrene concentration after adjustment for body mass index, physical activity level, urinary creatinine, age, and sex. The association of urinary 9-hydroxyphenanthrene and ambient phenantherene was significant in the crude model; however after adjustment for the abovementioned covariates, it was no more significant. We found significant correlations between exposure to ambient PM2.5-bounded PAHs and their urinary excretion. Considering the adverse health effects of PAHs in the pediatric age group, biomonitoring of PAHs should be underscored; preventive measures need to be intensified.

Urinary Trans, Trans-Muconic Acid is Not a Reliable Biomarker for Low-level Environmental and Occupational Benzene Exposures.

BACKGROUND: Benzene is a known occupational and environmental pollutant. Its urinary metabolite trans, trans-muconic acid (tt-MA) has been introduced by some environmental and occupational health regulatory associations as a biological index for the assessment of benzene exposure; however, recently, doubts have been raised about the specificity of tt-MA for low-level benzene exposures. In the present study, we investigated the association between urinary levels of tt-MA and inhalational exposure to benzene in different exposure groups. METHODS: Benzene exposure was assessed by personal air sampling. Collected benzene on charcoal tube was extracted by carbon disulfide and determined by a gas chromatograph (gas chromatography with a flame ionization detector). Urinary tt-MA was extracted by a strong anion-exchange column and determined with high-performance liquid chromatography-UV. RESULTS: Urinary levels of tt-MA in intensive benzene exposure groups (chemical workers and police officers) were significantly higher than other groups (urban and rural residents), but its levels in the last two groups with significant different exposure levels (mean = 0.081 ppm and 0.019 ppm, respectively) showed no significant difference (mean = 388 µg/g creatinine and 282 µg/g, respectively; p < 0.05). Before work shift, urine samples of workers and police officers showed a high amount of tt-MA and its levels in rural residents' samples were not zero. CONCLUSION: Our results suggest that tt-MA may not be a reliable biomarker for monitoring low-level (below 0.5 ppm) benzene exposures.

Primula auriculata Extracts Exert Cytotoxic and Apoptotic Effects against HT-29 Human Colon Adenocarcinoma Cells.

Primula auriculata (Tootia) is one of the most important local medicinal plants in Hamedan district, Iran. To investigate cytotoxicity and apoptosis induction of crude methanolic extract and different fraction of it, we compared several methods on HT-29 human colon Adenocarcinoma cells. Cancer cell proliferation was measured by 3-(4, 5­dimethylthiazolyl)2, 5­diphenyl­tetrazolium bromide (MTT) assay and apoptosis induction was analyzed by fluorescence microscopy (acridin orange/ethidium bromide, annexin V/propidium iodide staining, TUNEL assay and Caspase-3 activity assay). Crude methanolic extract (CM) inhibited the growth of malignant cells in a dose-dependent manner. Among solvent fractions, the dichloromethane fraction (CF) was found to be the most toxic compared to other fractions. With double staining methods, high percentage of 40 µg/mL of (CM) and (CF) treated cells exhibited typical characteristics of apoptotic cells. Apoptosis induction was also revealed by apoptotic fragmentation of nuclear DNA and activation of caspas-3 in treated cells. These findings indicate that crude methanolic extract and dichloromethan fraction of P.auriculata induced apoptosis and inhibited proliferation in colon cancer cells and could be used as a source for new lead structures in drug design to combat colon cancer.

Anticancer Activity a of Caspian Cobra (Naja naja oxiana) snake Venom in Human Cancer Cell Lines Via Induction of Apoptosis.

Cancer is the leading cause of death worldwide. Current anticancer drugs involve various toxic side effects; efforts are ongoing to develop new anticancer agents especially from the screening of natural compounds. Present study investigated cytotoxic effects and mode of cell death induced by the Caspian cobra venom in some human cancer cell lines. Cytotoxic effects of snake venom toxins (SVT) were investigated via monitoring of morphological changes, MTT, trypan blue exclusion and LDH release assays. Mechanism of cell death was determined by AO/EtBr double staining, caspase-3 activity assay, flow cytometric analysis of apoptosis and mitochondrial membrane potential measurement. In morphological analysis, apoptotic alterations related to apoptosis such as cytoplasmic blebbing, chromatin condensation and irregularity in shape were seen. IC50 of SVT in HepG2, MCF7and DU145 cell lines were 26.59, 28.85 and 21.17µg/mL, respectively and significantly different from the MDCK normal cell line (IC50=47.1 µg/mL). AO/EtBr double staining showed the best apoptotic/necrotic ratio at 15 µg/mL after 48 h. LDH release showed no significant differences between 10 µg/mL SVT and cisplatin. Flowcytometric analysis confirms mitochondrial membrane potential loss and more than 95% apoptotic cell death at 15 µg/mL. Caspase-3 was significantly activated at doses higher than 2.5 µg/mL with a maximal activity at 10 µg/mL. Results from this study demonstrate that SVT induces mitochondrial and caspase-3 dependent apoptosis in cancer cell lines with minimum effects on studied normal cell. This potential might candidate this venom as a suitable choice for cancer treatment.

Cobra venom cytotoxins; apoptotic or necrotic agents?

Organs homeostasis is controlled by a dynamic balance between cell proliferation and apoptosis. Failure to induction of apoptosis has been implicated in tumor development. Cytotoxin-I (CTX-I) and cytotoxin-II (CTX-II) are two physiologically active polypeptides found in Caspian cobra venom. Anticancer activity and mechanism of cell death induced by these toxins have been studied. The toxins were purified by different chromatographic steps and their cytotoxicity and pattern of cell death were determined by MTT, LDH release, acridine orange/ethidium bromide (AO/EtBr) double staining, flow cytometric analysis, caspase-3 activity and neutral red assays. The IC50 of CTX-II in MCF-7, HepG2, DU-145 and HL-60 was 4.1 ± 1.3, 21.2 ± 4.4, 9.4 ± 1.8 µg/mL and 16.3 ± 1.9 respectively while the IC50 of this toxin in normal MDCK cell line was 54.5 ± 3.9 µg/mL. LDH release suddenly increase after a specific toxins concentrations in all cell lines. AO/EtBr double staining, flow cytometric analysis and caspase-3 activity assay confirm dose and time-dependent induction of apoptosis by both toxins. CTX-I and CTX-II treated cells lost their lysosomal membrane integrity and couldn't uptake neutral red day. CTX-I and CTX-II showed significant anticancer activity with minimum effects on normal cells and better IC50 compared to current anticancer drug; cisplatin. They induce their apoptotic effect via lysosomal pathways and release of cathepsins to cytosol. These effects were seen in limited rage of toxins concentrations and pattern of cell death rapidly changes to necrosis by increase in toxin's concentration. In conclusion, significant apoptogenic effects of these toxins candidate them as a possible anticancer agent.

Anticancer Activity of Cobra Venom Polypeptide, Cytotoxin-II, against Human Breast Adenocarcinoma Cell Line (MCF-7) via the Induction of Apoptosis.

PURPOSE: Breast cancer is a significant health problem worldwide, accounting for a quarter of all cancer diagnoses in women. Current strategies for breast cancer treatment are not fully effective, and there is substantial interest in the identification of novel anticancer agents especially from natural products including toxins. Cytotoxins are polypeptides found in the venom of cobras and have various physiological effects. In the present study, the anticancer potential of cytotoxin-II against the human breast adenocarcinoma cell line (MCF-7) was investigated. METHODS: The cytotoxic effects of cytotoxin-II were determined by morphological analysis and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The mode and mechanism of cell death were investigated via acridine orange/ethidium bromide (AO/EtBr) double staining, flow cytometric analysis of cell death, detection of mitochondrial membrane potential, measurement of intracellular reactive oxygen species (ROS), annexin V/propidium iodide staining, and caspase-9 activity assays. RESULTS: The half maximal inhibitory concentration (IC50) of cytotoxin-II in MCF-7 cells was 4.18±1.23 µg/mL, while the value for cisplatin was approximately 28.02±1.87 µg/mL. Morphological analysis and AO/EtBr double staining showed typical manifestations of apoptotic cell death (in doses lower than 8 µg/mL). Dose- and time-dependent ROS generation, loss of mitochondrial membrane potential, caspase-9 activation, and cell cycle arrest were observed in their respective tests. CONCLUSION: In conclusion, cytotoxin-II has potent anticancer effects in the MCF-7 cell line, which are induced via the intrinsic pathways of apoptosis. Based on these findings, cytotoxin-II is a suitable choice for breast cancer treatment.