Gloria Barbosa-Sabanero

Exposure to organic solvents and cytogenetic damage in exfoliated cells of the buccal mucosa from shoe workers

PurposeWe determined the solvents mainly used in shoe making and their genotoxic effects.MethodsThirty-four exposed shoe workers and 34 unexposed control subjects, paired by age and sex, were compared. Occupational exposure was determined by using monitors 3M. Solvents were assessed by gas chromatography. Exfoliated buccal cells were obtained from each subject to determine the incidence of micronuclei and other nuclear abnormalities. One thousand cells were counted in each subject.ResultsSolvents detected were acetone, ethyl acetate, methyl ethyl ketone, and toluene. The incidence of nuclear abnormalities was significatively higher in the exposed group when compared to the control group. A positive relationship between the incidence of micronuclei and the toluene concentration in the environment was found.ConclusionsToluene shows an important genotoxic effect. As the micronuclei test is an effective, fast, simple and low cost biomarker to identify cytogenetic effects, we suggest its utilization as a preventive test of genotoxicity.

SUBCELLULAR LOCALIZATION OF THE NAD+-DEPENDENT ALCOHOL DEHYDROGENASE IN ENTAMOEBA HISTOLYTICA TROPHOZOITES

The protozoan parasite Entamoeba histolytica is an ancient eukaryotic cell that shows morphologically atypical organelles and differs metabolically from higher eukaryotic cells. The aim of this study was to determine the subcellular localization of ameba NAD+-dependent alcohol dehydrogenase (ADH2). The enzyme activity was present in soluble and mainly in particulate material whose density was 1.105 in a sucrose gradient. By differential centrifugation, most of the ADH activity sedimented at 160,000 g (160,000-g pellet), similar to the Escherichia coli polymeric ADHE. In the Coomassie staining of the 160,000-g pellet analyzed by electrophoresis, a 96-kDa protein was more prominent than in other fractions; this band was recognized by antibodies against Lactococcus lactis ADHE. By gold labeling, the antibodies recognized the granular material that mainly constitutes the 160,000-g pellet and a material that sedimented along with the internal membrane vesicles. By negative staining, the 160,000-g fraction showed helical rodlike structures with an average length of 103 nm; almost no membrane vesicles were observed in this pellet. In internal membrane fractions, no rodlike structures were found, but protomerlike round structures were observed. These results indicate that the main amebic NAD+-dependent ADH2 activity is naturally organized as rodlike helical particles, similar to bacterial ADHE. Detection of ADH2 in membrane fractions might be explained by cosedimentation of the multimeric ADH during membrane purification.

Arachidonic and oleic acid exert distinct effects on the DNA methylome.

ABSTRACT Abnormal fatty acid metabolism and availability are landmarks of metabolic diseases, which in turn are associated with aberrant DNA methylation profiles. To understand the role of fatty acids in disease epigenetics, we sought DNA methylation profiles specifically induced by arachidonic (AA) or oleic acid (OA) in cultured cells and compared those with published profiles of normal and diseased tissues. THP-1 monocytes were stimulated with AA or OA and analyzed using Infinium HumanMethylation450 BeadChip (Illumina) and Human Exon 1.0 ST array (Affymetrix). Data were corroborated in mouse embryonic fibroblasts. Comparisons with publicly available data were conducted by standard bioinformatics. AA and OA elicited a complex response marked by a general DNA hypermethylation and hypomethylation in the 1–200 μM range, respectively, with a maximal differential response at the 100 μM dose. The divergent response to AA and OA was prominent within the gene body of target genes, where it correlated positively with transcription. AA-induced DNA methylation profiles were similar to the corresponding profiles described for palmitic acid, atherosclerosis, diabetes, obesity, and autism, but relatively dissimilar from OA-induced profiles. Furthermore, human atherosclerosis grade-associated DNA methylation profiles were significantly enriched in AA-induced profiles. Biochemical evidence pointed to β-oxidation, PPAR-α, and sirtuin 1 as important mediators of AA-induced DNA methylation changes. In conclusion, AA and OA exert distinct effects on the DNA methylome. The observation that AA may contribute to shape the epigenome of important metabolic diseases, supports and expands current diet-based therapeutic and preventive efforts.

Ghrelin in small‐for‐gestational age (SGA) newborn babies: a cross‐sectional study

Background  Small‐for‐gestational newborn babies may have long‐term metabolic consequences. Among the main hormones possibly involved in foetal growth regulation are the IGFs, IGFBPs and the recently described ghrelin.

Detection of the presence of antibodies against Toxoplasma gondii in human colostrum by Raman spectroscopy and principal component analysis

More than 60 million people in the United States and 23 million people in Mexico probably are infected with the Toxoplasma parasite, but very few have symptoms because the immune system usually keeps the parasite from causing illness. However, for people whose immune system is compromised, the consequences can be fatal. Toxoplasmosis is detected indirectly by different serological tests, where the sample requires a previous preparation. We analyze the feasibility to use Raman spectroscopy and principal component analysis (PCA) as an alternative method to detect the presence or absence of antibodies IgG (immunoglobulin G), IgM (immunoglobulin M), and IgA (immunoglobulin A), against Toxoplasma gondii, in a simple and fast way, in samples of human colostrum from a group of volunteers who were in contact with the parasite and others who were not in contact with the parasite.

Comparative evaluation of three different ELISA assays and HPLC-ESI-ITMS/MS for the analysis of Nε-carboxymethyl lysine in food samples

Nε-carboxymethyl-lysine (CML) is measured in food, but there is a controversy concerning the most convenient yet reliable method(s) for this task. This work compares three different ELISA assays and HPLC-ESI-ITMS/MS for the analysis of CML in several food items. The four methods showed the same decreasing order of CML concentration: beef, bacon>chicken > fish>dairy products>grain products>fruits/vegetables. HPLC-ESI-ITMS/MS results highly correlated with those obtained by ELISA performed with monoclonal CML-antibody (β=0.98, p<0.0001) whereas My Bio Source® kit results were not correlated with those provided by Lamider®. Small differences of CML concentrations in food items prepared by different culinary treatment were clearly distinguished by HPLC-ESI-ITMS/MS, but could not always be detected by ELISA. This work demonstrates a reasonable relationship between CM determined by ELISA and HPLC-ESI-ITMS/MS and therefore supports the implementation of ELISA in food CML/AGEs screening.

Human Lung Cancer Cell Line A-549 ATCC Is Differentially Affected by Supranutritional Organic and Inorganic Selenium

The effects of organic and inorganic forms of selenium (Se) on human cells have been extensively studied for nutritional concentrations; however, to date, little is known about the potential toxicity at supranutritional levels. In the present study we determined the effects of sodium selenite (SSe) and selenomethionine (SeMet) on cell growth and intracellular structures in lung cancer cells exposed at Se concentrations between 0 and 3 mM. Our results showed that SSe affected cell growth more rapidly than SeMet (24 h and 48 h, resp.). After 24 h of cells exposure to 0.5, 1.5, and 3 mM SSe, cell growth was reduced by 10, 50, and 60%, as compared to controls. After 48 h, nuclear fragmentation was evident in cells exposed to SSe, suggesting an induction to cell death. In contrast, SeMet did not affect cell proliferation, and the cells were phenotypically similar to controls. Microtubules and microfilaments structures were also affected by both Se compounds, again SSe being more toxic than SeMet. To our knowledge, this is the first report on the differential effects of organic and inorganic Se in supranutritional levels in lung cancer cells.

11 beta-hydroxysteroid dehydrogenase 2 promoter methylation is associated with placental protein expression in small for gestational age newborns

Highlights11&bgr;‐HSD2 protein expression is lower in placentas of SGA newborns.Placental 11&bgr;‐HSD2 expression is directly correlated to birthweight.HSD11B2 promoter methylation is higher in SGA placentas.AGA and SGA placentas have no variation in global placental methylation.Reduced 11&bgr;‐HSD2 protein expression may be modulated by methylation of the HSD11B2 promoter. ABSTRACT Small for gestational age infants have greater risk of developing metabolic diseases in adult life. It has been suggested that low birth weight may result from glucocorticoid excess in utero, a key mechanism in fetal programming. The placental enzyme 11‐beta hydroxysteroid dehydrogenase type 2 (11&bgr;‐HSD2, HSD11B2 gene) acts as a barrier protecting the fetus from maternal corticosteroid deleterious effects. Low placental 11&bgr;‐HSD2 transcription and activity have been associated with low birth weight, yet the mechanism regulating its protein expression is not fully understood. In the present study we aimed to analyze 11&bgr;‐HSD2 protein expression in placentas of adequate and small for gestational age (AGA and SGA, respectively) newborns from healthy mothers, and to explore whether 11&bgr;‐HSD2 protein expression could be modulated by DNA methylation. 11&bgr;‐HSD2 protein levels were measured by western blot in placental biopsies from term AGA and SGA infants (n = 10 per group). DNA methylation was profiled both globally and in the HSD11B2 promoter by liquid chromatography with UV detection and methylation‐specific melting curve analysis, respectively. We found lower placental 11&bgr;‐HSD2 protein expression and higher HSD11B2 promoter methylation in SGA compared to AGA. Promoter methylation was inversely correlated with both protein expression and, importantly, birth weight. No changes in global placental methylation were found. In conclusion, lower 11&bgr;‐HSD2 protein expression is associated with higher HSD11B2 promoter methylation, correlating with birth weight in healthy pregnancy. Our data support the role of 11&bgr;‐HSD2 in determining birth weight, providing evidence of its regulation by epigenetic mechanisms, which may affect postnatal metabolic disease risk.

Omega 3 Fatty Acids Supplementation and Oxidative Stress in HIV-Seropositive Patients. A Clinical Trial.

HIV-seropositive patients show high incidence of coronary heart disease and oxidative stress has been described as relevant key in atherosclerosis development. The aim of this study was to assess the effect of omega 3 fatty acids on different markers of oxidative stress in HIV-seropositive patients. We performed a randomized parallel controlled clinical trial in The Instituto Mexicano del Seguro Social, a public health hospital. 70 HIV-seropositive patients aged 20 to 55 on clinical score A1, A2, B1 or B2 receiving highly active antiretroviral therapy (HAART) were studied. They were randomly assigned to receive omega 3 fatty acids 2.4 g (Zonelabs, Marblehead MA) or placebo for 6 months. At baseline and at the end of the study, anthropometric measurements, lipid profile, glucose and stress oxidative levels [nitric oxide catabolites, lipoperoxides (malondialdehyde plus 4-hydroxialkenals), and glutathione] were evaluated. Principal HAART therapy was EFV/TDF/FTC (55%) and AZT/3TC/EFV (15%) without difference between groups. Treatment with omega 3 fatty acids as compared with placebo decreased triglycerides (-0.32 vs. 0.54 mmol/L; p = 0.04), but oxidative stress markers were not different between groups. Trial Registration ClinicalTrials.gov NCT02041520

Mammalian cells exposed to ionizing radiation: Structural and biochemical aspects.

Acute or chronic exposure to ionizing radiation is a factor that may be hazardous to health. It has been reported that exposure to low doses of radiation (less than 50 mSv/year) and subsequently exposure to high doses produces greater effects in people. It has been reported that people who have been exposed to low doses of radiation (less than 50 mSv/year) and subsequently are exposed to high doses, have greater effects. However, at a molecular and biochemical level, it is an unknown alteration. This study, analyzes the susceptibility of a biological system (HeLa ATCC CCL-2 human cervix cancer cell line) to ionizing radiation (6 and 60 mSv/90 s). Our research considers multiple variables such as: total protein profile, mitochondrial metabolic activity (XTT assay), cell viability (Trypan blue exclusion assay), cytoskeleton (actin microfilaments), nuclei (DAPI), and genomic DNA. The results indicate, that cells exposed to ionizing radiation show structural alterations in nuclear phenotype and aneuploidy, further disruption in the tight junctions and consequently on the distribution of actin microfilaments. Similar alterations were observed in cells treated with a genotoxic agent (200 μM H2O2/1h). In conclusion, this multi-criteria assessment enables precise comparisons of the effects of radiation between various line cells. However, it is necessary to determine stress markers for integration of the effects of ionizing radiation.