Chronic Exposure to Arsenic and Fluoride Starting at Gestation Alters Liver Mitochondrial Protein Expression and Induces Early Onset of Liver Fibrosis in Male Mouse Offspring.

The presence of arsenic (As) and fluoride (F-) in drinking water is of concern due to the enormous number of individuals exposed to this condition worldwide. Studies in cultured cells and animal models have shown that As- or F-induced hepatotoxicity is primarily associated with redox disturbance and altered mitochondrial homeostasis. To explore the hepatotoxic effects of chronic combined exposure to As and F- in drinking water, pregnant CD-1 mice were exposed to 2 mg/L As (sodium arsenite) and/or 25 mg/L F- (sodium fluoride). The male offspring continued the exposure treatment up to 30 (P30) or 90 (P90) postnatal days. GSH levels, cysteine synthesis enzyme activities, and cysteine transporter levels were investigated in liver homogenates, as well as the expression of biomarkers of ferroptosis and mitochondrial biogenesis-related proteins. Serum transaminase levels and Hematoxylin-Eosin and Masson trichrome-stained liver tissue slices were examined. Combined exposure at P30 significantly reduced GSH levels and the mitochondrial transcription factor A (TFAM) expression while increasing lipid peroxidation, free Fe 2+, p53 expression, and serum ALT activity. At P90, the upregulation of cysteine uptake and synthesis was associated with a recovery of GSH levels. Nevertheless, the downregulation of TFAM continued and was now associated with a downstream inhibition of the expression of MT-CO2 and reduced levels of mtDNA and fibrotic liver damage. Our experimental approach using human-relevant doses gives evidence of the increased risk for early liver damage associated with elevated levels of As and F- in the diet during intrauterine and postnatal period.

Geographical approach analysis of the impact of air pollution on newborn intrauterine growth and cord blood DNA damage in Mexico City.

BACKGROUND: Few epidemiologic studies have focused on the specific source of ambient air pollution and adverse health effects in early life. Here, we investigated whether air pollutants from different emission sources were associated with decreased birth anthropometry parameters and increased DNA adduct formation in mother-child pairs residing in the Mexico City Metropolitan Area (MCMA). METHODS: This cross-sectional study included 190 pregnant women recruited during their last trimester of pregnancy from two hospitals at MCMA, and a Modeling Emissions Inventory (MEI) to calculate exposure to ambient air pollutants from different emissions sources (area, point, mobile, and natural) for two geographical buffers 250 and 750 m radii around the participants households. RESULTS: Contaminants were positively correlated with umbilical cord blood (UCB) adducts, but not with maternal blood (MB) adducts. PM10 emissions (area and point sources, overall emissions), PM2.5 (point sources), volatile organic compounds (VOC), total organic compounds (TOC) from point sources were positively correlated with UCB adducts. Air pollutants emitted from natural sources were correlated with a decrease in MB and UCB adducts. PM10 and PM2.5 were correlated (p < 0.05) with a decrease in birth weight (BW), birth length (BL) and gestational age at term (GA). In multivariate analyses adjusted for potential confounders, PM10 was associated with an increase in UCB adducts. PM10 and PM2.5 from overall emissions were associated with a decrease in BW, BL and GA at term. IMPACT: Results suggested higher susceptibility of newborns compared to mothers to damage related to ambient air pollution. PMs are associated with birth anthropometry parameters and DNA damage in adjusted models, highlighting the need for more strict regulation of PM emissions.

Early Neurotoxic Effects of Inorganic Arsenic Modulate Cortical GSH Levels Associated With the Activation of the Nrf2 and NFκB Pathways, Expression of Amino Acid Transporters and NMDA Receptors and the Production of Hydrogen Sulfide.

Exposure to toxic metals and metalloids is an important cause of preventable diseases worldwide. Inorganic arsenic (iAs) affects several organs and tissues, causing neurobehavioral alterations in the central nervous system (CNS) that might lead to neurodegeneration. In this work, we wanted to explore the time- and dose-related changes on glutathione (GSH) levels in several regions of the CNS, such as the cortex, striatum, hippocampus, and cerebellum, to identify the initial cellular changes associated to GSH depletion due to iAs exposure. Mice received a single intraperitoneal injection containing 5 or 14 mg/kg sodium arsenite. Animals were killed at 2, 6, and 24 h. Significant depletion of GSH levels was observed in the cortex at 2 and 6 h, while on the striatum, hippocampus, or cerebellum regions, no significant changes were observed. GSH depletion in the cortex was associated with the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) and nuclear factor kappa B (NFκB) pathways, which led to the upregulation of xCT, excitatory amino acid carrier 1 (EAAC1), glutamate/aspartate transporter (GLAST), and glial glutamate transporter 1 (GLT-1), and the activation of the transsulfuration pathways, which led to the overproduction of H2S in the cortex and increased levels of GSH in the cortex and cerebellum at 24 h. In the cortex, the N-methyl-D-aspartate (NMDA) receptor subunits NR2A and NR2B were also altered at 24 h. These early effects were not homogeneous among different brain regions and indicate early neurotoxic alterations in the cortex and cerebellum.

Systemic L-Buthionine -S-R-Sulfoximine Treatment Increases Plasma NGF and Upregulates L-cys/L-cys2 Transporter and γ-Glutamylcysteine Ligase mRNAs Through the NGF/TrkA/Akt/Nrf2 Pathway in the Striatum.

Glutathione (GSH) is the most abundant intracellular antioxidant. GSH depletion leads to oxidative stress and neuronal damage in the central nervous system (CNS). In mice, the acute systemic inhibition of GSH synthesis by L-buthionine-S-R-sulfoximine (BSO) triggers a protective response and a subsequent increase in the CNS GSH content. This response might be modulated by a peripheral increment of circulating nerve growth factor (NGF). NGF is an important activator of antioxidant pathways mediated by tropomyosin-related kinase receptor A (TrkA). Here, we report that peripheral administration of BSO increased plasma NGF levels. Additionally, BSO increased NGF levels and activated the NGF/TrkA/Akt pathway in striatal neurons. Moreover, the response in the striatum included an increased transcription of nrf2, gclm, lat1, eaac1, and xct, all of which are involved in antioxidant responses, and L-cys/L-cys2 and glutamate transporters. Using antibody against NGF confirmed that peripheral NGF activated the NGF/TrkA/Akt/Nrf2 pathway in the striatum and subsequently increased the transcription of gclm, nrf2, lat1, eaac1, and xct. These results provide evidence that the reduction of peripheral GSH pools increases peripheral NGF circulation that orchestrates a neuroprotective response in the CNS, at least in the striatum, through the NGF/TrkA/Akt/Nrf2 pathway.

Association of the Promoter Methylation and the rs12917 Polymorphism of MGMT with Formation of DNA Bulky Adducts and the Risk of Lung Cancer in Mexican Mestizo Population.

O6-Methylguanine-DNA methyltransferase (MGMT) is an enzyme that repairs the DNA damage caused by the tobacco habit, and low activity of this enzyme has been associated with a risk of lung cancer (LC). Our objective was to determine the association of the promoter methylation and the rs12917 polymorphism of MGMT with formation of DNA bulky adducts and the risk of LC in the Mexican Mestizo population. In this study are included 431 subjects. High-resolution melting analysis was used to determine the polymorphism MGMT rs12917 and methylation levels. DNA bulky adducts were determined by 32P-postlabeling. Our results showed that MGMT rs12917 and higher levels of methylation in the MGMT promoter are associated with the risk of LC. The levels of adducts are related with the phe/phe genotype and, only in the cases group, with the hypermethylation (>50%) of MGMT; however, this last association was not statistically significant.

Prenatal exposure to particulate matter and ozone: Bulky DNA adducts, plasma isoprostanes, allele risk variants, and neonate susceptibility in the Mexico City Metropolitan Area.

Mexico City's Metropolitan Area (MCMA) includes Mexico City and 60 municipalities of the neighbor states. Inhabitants are exposed to emissions from over five million vehicles and stationary sources of air pollutants such as particulate matter (PM) and ozone. MCMA PM contains elemental carbon and organic carbon (OC). OCs include polycyclic aromatic hydrocarbons (PAHs), many of which induce mutagenic and carcinogenic DNA adducts. Gestational exposure to air pollution has been associated with increased risk of intrauterine growth restriction, preterm birth or low birth weight risk, and PAH-DNA adducts. These effects also depend on the presence of risk alleles. We investigated the presence of bulky PAH-DNA adducts, plasma 8-iso-PGF2α (8-iso-prostaglandin F2α ) and risk allele variants in neonates cord blood and their non-smoking mothers' leucocytes from families that were living in a highly polluted area during 2014-2015. The presence of adducts was significantly associated with both PM2.5 and PM10 levels, mainly during the last trimester of gestation in both neonates and mothers, while the last month of pregnancy was significant for the association between ozone levels and maternal plasma 8-iso-PGF2α . Fetal CYP1B1*3 risk allele was associated with increased adduct levels in neonates while the presence of the maternal allele significantly reduced the levels of fetal adducts. Maternal NQO1*2 was associated with lower maternal levels of adducts. Our findings suggest the need to reduce actual PM limits in MCMA. We did not observe a clear association between PM and/or adduct levels and neonate weight, length, body mass index, Apgar or Capurro score. Environ. Mol. Mutagen. 60:428-442, 2019. © 2019 Wiley Periodicals, Inc.

Systemic L-buthionine-S-R-sulfoximine administration modulates glutathione homeostasis via NGF/TrkA and mTOR signaling in the cerebellum.

Glutathione (GSH) is an essential component of intracellular antioxidant systems that plays a primordial role in the protection of cells against oxidative stress, maintaining redox homeostasis and xenobiotic detoxification. GSH synthesis in the brain is limited by the availability of cysteine and glutamate. Cystine, the disulfide form of cysteine is transported into endothelial cells of the blood-brain barrier (BBB) and astrocytes via the system xc-, which is composed of xCT and the heavy chain of 4F2 cell surface antigen (4F2hc). Cystine is reduced inside the cells and the L-type amino acid transporter 1 (LAT1) transports cysteine from the endothelial cells into the brain, cysteine is transported into the neurons through the excitatory amino acid transporter 3 (EAAT3), also known as excitatory amino acid carrier 1 (EAAC1). The mechanistic/mammalian target of rapamycin (mTOR) and neurotrophins can activate signaling pathways that modulate amino acid transporters for GSH synthesis. The present study found that systemic L-buthionine-S-R-sulfoximine (BSO) administration selectively altered GSH homeostasis and EAAT3 levels in the mice cerebellum. Intraperitoneal treatment of mice with 6 mmol/kg of BSO depleted GSH and GSSG in the liver at 2 h of treatment. The cerebellum, but not other brain regions, exhibited a redox response. The mTOR and the neuronal growth factor (NGF)/tropomyosin receptor kinase A (TrkA) signaling pathways were activated and lead to an increase in the protein levels of the EAAT3 transporter, which was linked to an increase in the GSH/GSSG ratio and GSH concentration in the cerebellum at 0.5 and 2 h, respectively. Therefore, the cerebellum responds to peripheral GSH depletion via activation of the mTOR and NGF/TrkA pathways, which increase the transport of cysteine for GSH synthesis.

Induction of c-Jun by air particulate matter (PM10) of Mexico city: Participation of polycyclic aromatic hydrocarbons.

The carcinogenic potential of urban particulate matter (PM) has been partly attributed to polycyclic aromatic hydrocarbons (PAHs) content, which activates the aryl hydrocarbon receptor (AhR). Here we report the effect of PM with an aerodynamic size of 10 µm (PM10) on the induction of AhR pathway in A549 cells, evaluating its downstream targets CYP1B1, IL-6, IL-8 and c-Jun. Significant increases in CYP1B1 protein and enzyme activity; IL-6 and IL-8 secretion and c-Jun protein were found in response to PM10. The formation of PAH-DNA adducts was also detected. The involvement of AhR pathway was confirmed with Resveratrol as AhR antagonist, which reversed CYP1B1 and c-Jun induction. Nevertheless, in IL-6 and IL-8 secretion, the Resveratrol was ineffective, suggesting an effect independent of this pathway. Considering the role of c-Jun in oncogenesis, its induction by PM may be contributing to its carcinogenic potential through induction of AhR pathway by PAHs present in PM10.

The GSTM1null (deletion) and MGMT84 rs12917 (Phe/Phe) haplotype are associated with bulky DNA adduct levels in human leukocytes.

Tobacco smoke and air pollutants contain carcinogens, such as polycyclic aromatic hydrocarbons (PAHs) and tobacco specific nitrosamines (TSNA), that are substrates of metabolizing enzymes generating reactive metabolites that can bind to DNA. Variation in the activity of these enzymes may modify the extent to which these metabolites can interact with DNA. We compared the levels of bulky DNA adducts in blood leukocytes from 93 volunteers living in Mexico City with the presence of 13 single nucleotide polymorphisms (SNPs) in genes related to PAH and TSNA metabolism (AhR rs2044853, CYP1A1 rs1048943, CYP1A1 rs1048943, CYP1A1 rs1799814, EPHX1 rs1051740, EPHX1 rs2234922, GSTM1 null, GSTT1 null and GSTP1 rs947894), DNA repair (XRCC1 rs25487, ERCC2 rs13181 and MGMT rs12917) and cell cycle (TP53 rs1042522). (32)P-postlabeling analysis was used to quantify bulky DNA adduct formation. Genotyping was performed using PCR-RFLP. The mean levels of bulky DNA adducts were 8.51±3.66 adducts/10(8) nucleotides (nt) in smokers and 8.38±3.59 adducts/10(8) nt in non-smokers, being the difference not statistically significant. Without taking into account the smoking status, GSTM1 null individuals had a marginally significant lower adduct levels compared with GSTM1 volunteers (p=0.0433) and individuals heterozygous for MGMT Leu/Phe had a higher level of bulky adducts than those who were homozygous wild type (p=0.0170). A multiple regression analysis model showed a significant association between the GSTM1 (deletion) and MGMT rs12917 (Phe/Phe) haplotype and the formation of DNA adducts in smokers (R(2)=0.2401, p=0.0215). The presence of these variants conferred a greater risk for higher adduct levels in this Mexican population.

Bioconversion of Agave tequilana fructans by exo-inulinases from indigenous Aspergillus niger CH-A-2010 enhances ethanol production from raw Agave tequilana juice.

Agave tequilana fructans are the source of fermentable sugars for the production of tequila. Fructans are processed by acid hydrolysis or by cooking in ovens at high temperature. Enzymatic hydrolysis is considered an alternative for the bioconversion of fructans. We previously described the isolation of Aspergillus niger CH-A-2010, an indigenous strain that produces extracellular inulinases. Here we evaluated the potential application of A. niger CH-A-2010 inulinases for the bioconversion of A. tequilana fructans, and its impact on the production of ethanol. Inulinases were analyzed by Western blotting and thin layer chromatography. Optimal pH and temperature conditions for inulinase activity were determined. The efficiency of A. niger CH-A-2010 inulinases was compared with commercial enzymes and with acid hydrolysis. The hydrolysates obtained were subsequently fermented by Saccharomyces cerevisiae to determine the efficiency of ethanol production. Results indicate that A. niger CH-A-2010 predominantly produces an exo-inulinase activity. Optimal inulinase activity occurred at pH 5.0 and 50 °C. Hydrolysis of raw agave juice by CH-A-2010 inulinases yielded 33.5 g/l reducing sugars, compared with 27.3 g/l by Fructozyme(®) (Novozymes Corp, Bagsværd, Denmark) and 29.4 g/l by acid hydrolysis. After fermentation of hydrolysates, we observed that the conversion efficiency of sugars into ethanol was 97.5 % of the theoretical ethanol yield for enzymatically degraded agave juice, compared to 83.8 % for acid-hydrolyzed juice. These observations indicate that fructans from raw Agave tequilana juice can be efficiently hydrolyzed by using A. niger CH-A-2010 inulinases, and that this procedure impacts positively on the production of ethanol.

Antineoplastic effect of iodine and iodide in dimethylbenz[a]anthracene-induced mammary tumors: association between lactoperoxidase and estrogen-adduct production.

Several groups, including ours, have reported that iodine exhibited antiproliferative and apoptotic effects in various cancer cells only if this element is supplemented as molecular iodine, or as iodide, to cells that are able to oxidize it with the enzyme thyroperoxidase. In this study, we analyzed the effect of various concentrations of iodine and/or iodide in the dimethylbenz[a]anthracene (DMBA) mammary cancer model in rats. The results show that 0.1% iodine or iodide increases the expression of peroxisome proliferator-activated receptor type γ (PPARγ), triggering caspase-mediated apoptosis pathways in damaged mammary tissue (DMBA-treated mammary gland) as well as in frank mammary tumors, but not in normal mammary gland. DMBA treatment induces the expression of lactoperoxidase, which participates in the antineoplastic effect of iodide and could be involved in the pro-neoplastic effect of estrogens, increasing the formation of DNA adducts. In conclusion, our results show that a supplement of 0.1% molecular iodine/potassium iodide (0.05/0.05%) exert antineoplastic effects, preventing estrogen-induced DNA adducts and inducing apoptosis through PPARγ/caspases in pre-cancer and cancerous cells. Since this iodine concentration does not modify the cytology (histology, apoptosis rate) or physiology (triiodothyronine and thyrotropin) of the thyroid gland, we propose that it be considered as an adjuvant treatment for premenopausal mammary cancer.

Arsenic species, AS3MT amount, and AS3MT gene expression in different brain regions of mouse exposed to arsenite.

Human exposure to inorganic arsenic (iAs) has been associated with cancer and serious injury to various internal organs, as well as peripheral neuropathy, endocrine disruption and diverse effects in the central nervous system (CNS). Using rodent models, it is possible to demonstrate As accumulation in the brain that leads to defects in operant learning, behavioral changes, and affect pituitary gonadotrophins. iAs biomethylation in the CNS is a significant process, yielding products that are more reactive and toxic than the parent compound. Mice received 2.5, 5, and 10 mg/kg/day sodium arsenite orally for 9 days. We investigated the distribution of iAs and its metabolites as well as the mRNA and protein expression of arsenic (III) methyltransferase (AS3MT), which encodes the key enzyme in iAs metabolism, in the cerebral cortex, hippocampus, striatum, mesencephalon, thalamus, cerebellum, hypothalamus, pons, medulla oblongata, and pituitary of mouse brain. Our findings show that methylated As metabolites are present in all brain regions studied suggesting that AS3MT is ubiquitously expressed in the brain and it is not inducible by dose of arsenite. There is also a dose-related accumulation of As species in all brain regions, with the highest accumulation observed in the pituitary. The higher distribution of arsenicals in pituitary can help to explain the neuroendocrine effects associated with iAs exposure.

Streptomyces bangladeshensis sp. nov., isolated from soil, which produces bis-(2-ethylhexyl)phthalate.

The taxonomic position of an actinomycete strain isolated from soil from Natore, Bangladesh, was examined by using a polyphasic approach. The strain, designated AAB-4(T), was assigned to the genus Streptomyces on the basis of chemical and morphological criteria. It formed Rectiflexibiles aerial hyphae that carried long chains of rounded spores. The 16S rRNA gene of strain AAB-4(T) was sequenced directly and then compared with those of previously studied streptomycetes following the generation of two phylogenetic trees by using maximum-likelihood and neighbour-joining algorithms. This confirmed the assignment of the novel strain to the genus Streptomyces. This strain showed a high level of 16S rRNA gene sequence similarity to Streptomyces thermoviolaceus, Streptomyces thermodiastaticus and Streptomyces longisporus, among others, but could be distinguished from them by phenotypic and physiological traits. This micro-organism produces bis-(2-ethylhexyl)phthalate, an antibacterial and antifungal agent. It is proposed that strain AAB-4(T) be classified as a novel species within the genus Streptomyces, as Streptomyces bangladeshensis sp. nov. (type strain, AAB-4(T)=LMG 22738(T)=NRRL B-24326(T)).

Streptomyces mexicanus sp. nov., a xylanolytic micro-organism isolated from soil.

The taxonomic position of a thermophilic actinomycete strain isolated from soil was examined using a polyphasic approach. The strain, designated CH-M-1035T, was assigned to the genus Streptomyces on the basis of chemical and morphological criteria. It formed Rectiflexibiles aerial hyphae that carried long chains of rounded, smooth spores. The almost complete nucleotide sequence of the 16S rRNA gene of strain CH-M-1035T was determined and its comparison with the 16S rDNA sequences of previously studied streptomycetes confirmed the assignment of the novel strain to the genus Streptomyces. Strain CH-M-1035T clustered with species belonging to the Streptomyces thermodiastaticus clade in the 1 6S-rDNA-based phylogenetic tree. However, the phenotypic properties of strain CH-M-1035T differed from those of the recognized species within this clade. Therefore, it is proposed that strain CH-M-1035T be classified as a novel species within the genus Streptomyces, as Streptomyces mexicanus (type strain CH-M-1035T =DSM 41796T =BM-B-384T =NRRL B-24196T).

Genes and enzymes of the acetyl cycle of arginine biosynthesis in Corynebacterium glutamicum: enzyme evolution in the early steps of the arginine pathway.

A cluster of arginine biosynthetic genes of Corynebacterium glutamicum ATCC 13032, comprising argJ, argB and argD as well as part of argC and argF, has been cloned by heterologous complementation of an Escherichia coli argE mutant. The gene order has been established as argCJBDF by sequencing the entire 4.4 kb cloned DNA fragment. The C. glutamicum argB gene can be transcribed in E. coli cells from an internal promoter located in the coding part of the preceding argJ gene, whereas transcription of the argJ gene appears vector-dependent. Expression of the corynebacterial argB gene is repressed by arginine in the native host but not in recombinant E. coli cells. Feedback inhibition of the corresponding N-acetylglutamate kinase activity was observed both in cell extracts of C. glutamicum and in recombinant E. coli argB auxotrophic strains. Extracts of E. coli cells carrying cloned corynebacterial DNA display an ornithine acetyltransferase activity (encoded by argJ) which alleviates the acetylornithinase (encoded by argE) deficiency of the enterobacterial host. In contrast to Bacillus stearothermophilus ornithine acetyltransferase which also exhibits acetylglutamate synthase activity, C. glutamicum ornithine acetyltransferase appears monofunctional. ArgA and ArgB proteins from different sources share highly significant similarities. The evolutionary implications of these data are discussed.