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The presence of mycotoxins in cereals and cereal products remains a significant issue. The use of natural ingredients such as pumpkin and whey, which contain bioactive compounds, could be a strategy to reduce the use of conventional chemical preservatives. The aim of the present work was to study the bioaccessibility of aflatoxin B1 (AFB1) and ochratoxin (OTA) in bread, as well as to evaluate the effect of milk whey (with and without lactic acid bacteria fermentation) and pumpkin on reducing mycotoxins bioaccessibility. Different bread typologies were prepared and subjected to an in vitro digestion model. Gastric and intestinal extracts were analyzed by HPLC–MS/qTOF and mycotoxins bioaccessibility was calculated. All the tested ingredients but one significantly reduced mycotoxin intestinal bioaccessibility. Pumpkin powder demonstrated to be the most effective ingredient showing significant reductions of AFB1 and OTA bioaccessibility up to 74% and 34%, respectively. Whey, fermented whey, and the combination of pumpkin-fermented whey showed intestinal bioaccessibility reductions between 57–68% for AFB1, and between 11–20% for OTA. These results pointed to pumpkin and milk whey as potential bioactive ingredients that may have promising applications in the bakery industry.

Microbial fermentation with lactic acid bacteria (LAB) is a natural food biopreservation method. Yellow mustard and milk whey are optimum substrates for LAB fermentation. The aim of the present study was to evaluate the bioaccessibility and bioavailability of bioactive compounds from yellow mustard flour and milk whey both with and without LAB fermentation. All extracts were subjected to a simulated digestion process. Total polyphenols, DL-3-phenyllactic acid (PLA), lactic acid, and the antioxidant activity were determined in the studied matrices before and after simulated digestion. Yellow mustard flour was significantly richer in total polyphenols, whereas significantly higher concentrations of PLA and lactic acid were observed in milk whey. Similar antioxidant activity was determined in both ingredients being in all cases strongly reduced after in vitro digestion. Higher bioaccessibility was found for polyphenols and PLA in milk whey. Transepithelial transport of total polyphenols was higher in yellow mustard flour compared to milk whey, reaching bioavailability values between 3-7% and 1-2%, respectively. PLA transepithelial transport was only significant in both fermented matrices with bioavailability around 4-6%. Transepithelial transport of lactic acid reached values of 31-34% (bioavailability ∼ 22%) and 15-78% (bioavailability ∼ 3%) in milk whey and yellow mustard flour, respectively. LAB fermentation showed beneficial effects on enriching extracts with PLA, lactic acid, and antioxidant activity, as well as increasing bioaccessibility of these acids in yellow mustard flour and total polyphenol bioavailability in milk whey. Results pointed to yellow mustard flour and milk whey as natural preservative ingredients used in the food industry, especially when fermented with LAB.

Enniatins (ENs) are depsipeptide mycotoxins produced by Fusarium fungi. They are known for their capacity to modulate cell membrane permeability and disruption of ionic gradients, affecting cell homeostasis and initiating oxidative stress mechanisms. The effect of the acute toxicity of ENs A, A1, B and B1 at two different concentrations after 8 h of exposure was analysed in Wistar rats by a transcriptional approach. The following key mitochondrial and nuclear codified genes related to the electron transport chain were considered for gene expression analysis in stomach, liver, kidney and lower intestine by quantitative Real-Time PCR: mitochondrially encoded NADH dehydrogenase 1 (MT-ND1), mitochondrially encoded cytochrome c oxidase 1 (MT-COX1), succinate dehydrogenase flavoprotein subunit A and ATP synthase F1 subunit alpha, respectively. Moreover, the expression of markers involved in oxidative stresssuperoxide dismutase 1 (SOD1), glutathione peroxidase 1 (Gpx1), heme oxygenase 1, apoptosis B-cell lymphoma 2, Bcl2 Associated protein X (Bax), tumor suppressor protein (p53), inhibition of apoptosis nuclear factor kappa of activated B cells, immune system interleukin 1β and intestinal tight junction Occludin merely in lower intestine tissues have been investigated. For mitochondrial genes, the main differences were observed for MT-ND1 and MT-COX1, showing its deficiency in all selected organs. With regard to the intestinal barrier’s cellular response to oxidative stress, the activity of the antioxidant gene SOD1 was decreased in a dose-dependent manner. Similarly, the catalytic enzyme GPx1 was also downregulated though merely at medium dose employed. On the contrary, the pro-apoptotic Bax and p53 regulators were activated after ENs exposure, reporting a significant increase in their expression. Furthermore, the alteration of intestinal permeability was assessed by the abnormal activity of the tight junction protein occludin. In summary, ENs may generate mitochondrial disorders and induce oxidative stress in intestinal barrier function.

Introduction In clinical practice, celiac disease (CD) is monitored through anti-transglutaminase (TGA-IgA) antibody levels. The normalization of serum levels in successive periodic measurements indicates good response and adherence to dietary treatment. Objectives To evaluate the factors associated with the evolution of TGA-IgA antibodies and their association with dietary non-compliance and diseases related to CD. Methods This prospective observational study was carried out in 254 participants, who were recruited from patients from a hospital in southern Spain. Information about sex, age, serological test results, HLA DQ2/DQ8 haplotypes, mucosal atrophy, gastrointestinal and extra-intestinal symptoms, as well as diagnosis of diseases related to CD, was collected. Results Clinical manifestations, such as diarrhoea, abdominal pain and weight loss, showed differences according to sex and age. Children under 18 years of age presented a degree of total or severe atrophy of the intestinal villi. TGA-IgA antibodies concentrations were directly associated with the number of digestive disorders manifested by the patient and the record of dietary non-compliance and inversely related to the number of extra-digestive disorders. Conclusions Adolescents between 12 and 18 years old were the least monitored as well as the group with more extra-intestinal symptoms reported. Therefore, it is necessary to develop strategies in clinical practice aimed at this population group and continuous monitoring should be implemented to improve life quality and reduce complications that may arise in the long term., Celiac disease, TGA-IgA antibodies, Diet, Dietary non-compliance

Food and feed are daily exposed to mycotoxin contamination which effects may be counteracted by antioxidants like carotenoids. Some mycotoxins as well as carotenoids penetrate the blood brain barrier (BBB) inducing alterations related to redox balance in the mitochondria. Therefore, the in vitro BBB model ECV304 was subcultured for 7 days and exposed to beauvericine, enniatins, ochratoxin A, zearalenone (100 nM each), individually and combined, and pumpkin extract (500 nM). Reactive oxygen species were measured by fluorescence using the dichlorofluorescein diacetate probe at 0 h, 2 h and 4 h. Intracellular ROS generation reported was condition dependent. RNA extraction was performed and gene expression was analyzed by qPCR after 2 h exposure. The selected genes were related to the Electron Transport Chain (ETC) and mitochondrial activity. Gene expression reported upregulation for exposures including mycotoxins plus pumpkin extract versus individual mycotoxins. Beauvericin and Beauvericin-Enniatins exposure significantly downregulated Complex I and pumpkin addition reverted the effect upregulating Complex I. Complex IV was the most downregulated structure of the ETC. Thioredoxin Interacting Protein was the most upregulated gene. These data confirm that mitochondrial processes in the BBB could be compromised by mycotoxin exposure and damage could be modulated by dietary antioxidants like carotenoids.

Introducción: La enfermedad celíaca (EC) se caracteriza por una gran variedad de signos, síntomas y enfermedades asociadas en su forma de presentación, incluso puede cursar de forma asintomática. Recientes estudios muestran la variación del espectro clínico según la edad. En niños y niñas pequeñas predomina la forma clásica con síntomas como distensión abdominal, disminución del apetito, diarrea y pérdida de peso. Las manifestaciones atípicas frecuentes en niños mayores son dolor abdominal, estreñimiento, reflujo, vómitos, fatiga, talla baja. En adultos se observa una reducción de la forma de presentación clásica e incremento de la no clásica.Material y métodos: Se realizó una revisión sistemática en la que se han incluido los artículos sobre signos y síntomas de la EC clásica y atípica en niños y adultos, además de las enfermedades que se asocian de forma frecuente a esta patología. Las bases de datos utilizadas para la búsqueda de los artículos fueron PubMed, Web of Science y Scopus, de los últimos seis años (2014-2019). Se han evaluado un total de 164 artículos tras los criterios de selección, de los cuales se han incluido en el estudio los 20 artículos más relevantes.Resultados: La EC se diagnostica con mayor frecuencia en mujeres y a edades más tempranas que a los hombres. En niños menores de dos años predomina la EC clásica, aunque de forma más intensa que en niños y niñas pequeñas. En niños y niñas mayores, adolescentes y edad adulta varía la forma de presentación hacia la atípica o no clásica y asintomática.Conclusiones: El incremento en la prevalencia en los últimos años, gran heterogeneidad de síntomas, enfermedades asociadas y la variación del espectro clínico hacia una forma atípica con síntomas extraintestinales causa retraso en su diagnóstico por lo que deben reconocerse para que se detecte de manera más precoz.

Several mycotoxins exert their effect on the immunological system; some are classified as immunotoxic. Jurkat T-cells were used to study toxic effects of beauvericin (BEA) and enniatin B (ENN B). Both are not legislated mycotoxins with increasing presence in feed and food. Concentrations studied were from 1 to 15 μM at 24, 48 and 72 h. Cell death by increasing the percentage of apoptotic/necrotic cells was: BEA > ENN B. IC50 values ranged from 3 to 7.5 μM for BEA. ENN B 15 μM decreased viability (21-29%). The percentage of apoptotic/necrotic cells was BEA > ENN B at 24 h but not at 48 h. Caspase-3&7 activation profile varied, although both mycotoxins increased this activation. No difference in ROS production for any mycotoxin was observed. Arrest in S phase for both mycotoxins was obtained. BEA increased the percentage of DNA in the tail (18% and 20%) with respect to the control, whereas not for ENN B. In summary, cytotoxicity of BEA involved mitochondrial alterations; while ENN B only at highest concentrations and time assayed. BEA had cell cycle disturbances and apoptotic and apoptotic/necrotic cells increased; for ENN B these were not evident. Different toxic responses in Jurkat T-cells may be involved in BEA and ENN B toxicity.

Nonceliac gluten sensitivity (NCGS) is a gluten-related gastrointestinal disorder distinct from celiac disease (CD) and gluten allergy that is not easy to diagnose due to the lack of biomarkers. It is characterized by intestinal symptoms and extraintestinal manifestations with the consumption of gluten-containing foods. In contrast to CD, NCGS patients do not present a genetic predisposition or intestinal villi atrophy. Recent studies question the proinflammatory triggering activity of α-gliadin fraction contained in wheat, since it has been demonstrated that the amylase-trypsin inhibitors (ATIs) exert a strong activating effect on the innate immune response. We aimed to analyze the role of ATIs in the activation of innate immunity and in the development of the symptoms characteristic of NCGS. A systematic literature search was made using databases such as MEDLINE, SciELO, Science Direct, and Scopus, with focus on key words such as "amylase-trypsin inhibitors," "wheat," "gluten," and "celiac." Many studies are available on the structure, inhibition mechanism, and immune system effects of ATIs, mainly focused on IgE-mediated reactions. Recently, with the increase of NCGS interest, has increased the literature on the capacity of ATIs contained in wheat to activate the innate immune system. Literature published to date questions the relationship between activation of the innate immune system and gluten in NCGS. ATIs may have acted as interfering contaminant of gluten and appear as potential activator of innate immunity in NCGS patients. In view of their potential impact, more interventional studies are needed to demonstrate the proinflammatory effect of ATIs.

C. maxima (var. Delica), a variety of pumpkin, is well known for its high concentration on carotenoids, possessing dietary benefits and antioxidant properties. Aflatoxins and enniatins are common mycotoxins present in food and feed with an extended toxicity profile in humans and animals. Both types of substances reach a wide range of tissues and organs and have the capability to penetrate the blood brain barrier. Since carotenoids and mycotoxins have been reported to modify diverse mitochondrial processes individually, transcriptional in vitro studies on human epithelial cells ECV 304 were conducted to analyze the relative expression of 13 mitochondria related genes. ECV 304 cells were differentiated for 9 days and treated for 2 h with: a) pumpkin (500 nM); b) aflatoxins (100 nM); c) enniatins (100 nM); d) aflatoxins (100 nM) and pumpkin (500 nM); e) enniatins (100 nM) and pumpkin (500 nM). Even at low concentrations, dietary carotenoids activity on mitochondrial genes expression reported a beneficial effect and, for most of the genes studied across the Electron Transport Chain (ETC), developed a protective effect when mixed with aflatoxins (AFs) or enniatins (ENs).

Mycotoxins are considered to be a major risk factor affecting human and animal health as they are one of the most dangerous contaminants of food and feed. This review aims to compile the research developed up to date on the toxicological effects that mycotoxins can induce on human health, through the examination of a selected number of studies in vivo. AFB1 shows to be currently the most studied mycotoxin in vivo, followed by DON, ZEA and OTA. Scarce data was found for FBs, PAT, CIT, AOH and Fusarium emerging mycotoxins. The majority of them concerned the investigation of immunotoxicity, whereas the rest consisted in the study of genotoxicity, oxidative stress, hepatotoxicity, cytotoxicity, teratogenicity and neurotoxicity. In order to assess the risk, a wide range of different techniques have been employed across the reviewed studies: qPCR, ELISA, IHC, WB, LC-MS/MS, microscopy, enzymatic assays, microarray and RNA-Seq. In the last decade, the attention has been drawn to immunologic and transcriptomic aspects of mycotoxins' action, confirming their toxicity at molecular level. Even though, more in vivo studies are needed to further investigate their mechanism of action on human health.

Enniatins (ENs) are emerging mycotoxins produced by Fusarium fungi which are cytotoxic also at low concentrations due to its ionophoric properties. The aim of this study was to evaluate the hepatic toxicity of ENs exposure at different concentrations in Wistar rats through a proteomic approach. Animals were intoxicated by oral gavage with medium (EN A 256, ENA1 353, ENB 540, ENB1 296 μg/mL) and high concentrations (ENA 513, ENA1 706, ENB 1021, ENB1 593 μg/mL) of an ENs mixture and sacrificed after 8 h. Protein extraction was performed using powdered liver. Peptides were analyzed using a liquid chromatography coupled with a quadrupole time-of-flight mass spectrometer. Proteins were filtered by abundance using Mass Professional Profiler software (Agilent Technologies) and 57 were differentially expressed when compared to the control. In terms of abundance, the liver biomarker Carboamoyl-phosphate synthase showed the highest levels in all conditions employed while actin-1 had the lowest. Bioinformatic analysis using DAVID platform reported acetylation, nucleotide phosphate-binding region:NAD and catalytic activity as the most represented terms. Furthermore, metabolism was the most significant and enriched pathway in Reactome overrepresentation. In conclusion, ENs acute exposure caused protein expression changes related to major cellular processes in rats, hinting its involvement in liver disturbance.

Hepatotoxicity of three Fusarium mycotoxins, beauvericin (BEA) and two enniatins (ENNs) ENN A1 and ENN B1, in hepatocarcinoma cells (HepG2) were evaluated and compared. Concentrations used were 1.5 and 3 μM at 24, 48 and 72 h for each mycotoxin. Flow cytometry was used to examine enniatins effects on cell proliferation, to characterize the cell cycle phase where the cells blocked and to study the mitochondria role in ENNs-induced apoptosis. ENN B1 treated cells showed a time dependent G1 blockade at both concentrations used. ENN A1 and BEA decreased the apoptotic-necrotic percentage of cells comparing to control and disrupted the MMP as observed by TMRM and ToPro-3 fluorochromes signal. It is proposed a decreasing mycotoxin order by number of effects as follows: BEA > ENN B1 > ENN A1, with 47, 20 and 16%, respectively out of all situations compared.

The aim of this study was to investigate the influence of prebiotic compounds (cellulose and inulin), food ingredients (milk whey, β-lactoglobulin and calcium caseinate) and several probiotic microorganisms on the bioaccessibility of beauvericin (BEA), enniatins (ENs A, A1, B, B1), deoxynivalenol (DON) and zearalenone (ZEA) present in wheat crispy bread produced with wheat flour previously fermented with F. tricinctum, F. culmorum and G. zeae. The bioaccessibility of mycotoxins was determined by a dynamic simulated gastrointestinal digestion system, imitating the human digestive physiological conditions of the gastrointestinal tract. Mycotoxins were determined in the simulated intestinal fluids by liquid chromatography-tandem mass spectrometry (LC-MS/MS). EN bioaccessibility ranged from 15.1 to 30.6%, whereas the values evidenced for BEA ranged from 12 to 19%. DON showed bioaccessibility data ranging from 0.8 to 5.6% whereas for ZEA the data evidenced ranged from 26 to 44%. The bioaccessibility reduction evidenced using probiotic microorganisms for the mycotoxins studied ranged from 21 to 27.1% for ENs, from 29 to 39.7% for DON, from 41 to 57% for ZEA and from 6.6 to 10.5% for BEA. The addition of prebiotic and bioactive microorganisms decreased the bioaccessibility of mycotoxins, with a concentration-dependent behavior, thus being a potential strategy for reducing human exposure to these minor mycotoxins.

Beauvericin (BEA) is a bioactive compound produced by the secondary metabolism of several Fusarium strains and known to have various biological activities. This study investigates the influence of several dietary fibers (galactomanan, glucomannan, citrus fiber, bamboo fiber, carrot fiber, pie fiber, β-glucan, xilan, and cellulose) and probiotic strains ( Lactobacillus animalis , Lb. casei , Lb. casei , Lb. plantarum , Lb. rhuminis , Lb. casei casei , Bifidobacterium breve , Bf. Adolescents , Bf. bifidum , Corynebacterium vitaeruminis , Streptococcus faecalis , Eubacterium crispatus , and Saccharomyces cerevisiae ) on the minor Fusarium mycotoxin BEA bioaccessibility employing a model solution. The bioaccessibility was determined using a simulated gastrointestinal digestion that mimics the physiological conditions of the digestive tract until the colonic compartment. The determination of BEA in the intestinal fluids was carried out by liquid chromatography–mass spectrometry detection (LC–MS). The reduction of BEA bioaccessibility in the experiments carried out using the prebiotic compounds ranged from 60 to 80%, whereas in the trials carried out using the probiotic strains the bioaccessibility observed ranged from 30 to 85%. A BEA degradation product produced by colonic fermentation was identified using the technique of LC–MS-LIT.

Fungi producers of mycotoxins are able to synthesize more than one toxin. Alternariol (AOH) is one of the mycotoxins produced by several Alternaria species, the most common one being Alternaria alternata. The toxins 3-Acetyl-deoxynivalenol (3-ADON) and 15-Acetyl-deoxynivalenol (15-ADON) are acetylated forms of deoxynivalenol (DON) produced by Fusarium graminearum. In the present work it is determined and evaluated the toxic effects of binary and tertiary combination treatment of HepG2 cells with AOH, 3-ADON and 15-ADON, by using the MTT assay (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide), to subsequently apply the isobologram method and elucidate if the mixtures of these mycotoxins produced synergism, antagonism or additive effect; and lastly, to analyze mycotoxins conversion into metabolites produced and released by HepG2 cells after applying the treatment conditions by liquid chromatography tandem mass spectrometry (LC-MS/MS) equipment and extracted from culture media. HepG2 cells were treated at different concentrations over 24, 48 and 72h. IC50 values detected at all times assayed, ranged from 0.8 to >25μM in binary combinations; while in tertiary it ranged from 7.5 to 12μM. Synergistic, antagonism or additive effect detected in the mixtures of these mycotoxins was different depending on low or high concentration. Among all four mycotoxins combinations assayed, 15-ADON+3-ADON presented the highest toxic potential. At all assayed times, recoveries values oscillated depending on the time and combination studied.

espanolIntroduccion: suelen usarse diversos indices dieteticos para medir la calidad de la nutricion en edades avanzadas. Sin embargo, ninguno de ellos fue disenado para evaluar la nutricion con el objetivo de evitar discapacidades en la poblacion anciana. Objetivos: identificar en la literatura cientifica los “nutrientes e ingestas” involucrados en el envejecimiento, asi como proponer un nuevo indice, considerando esta informacion, que sirva para evaluar la calidad de la nutricion con objeto de prevenir enfermedades relacionadas con el envejecimiento. Metodos: se realizo una revision bibliografica, obteniendo informacion sobre los nutrientes asociados con el envejecimiento. Todos estos nutrientes fueron considerados en el diseno de un nuevo Indice de Dieta de Envejecimiento Saludable (HADI). A continuacion, se realizo un estudio transversal con dos muestras de ancianos (muestreo de conveniencia), recogiendo datos nutricionales y dieteticos, calculando las puntuaciones de diferentes indices dieteticos validados y comparandolos con HADI para validar los resultados. Resultados: cuarenta y ocho manuscritos fueron finalmente utilizados para el analisis de texto completo. Se encontraron asociaciones entre enfermedades cardiovasculares y macronutrientes, fibra dietetica, sodio y vitamina D; cancer y acidos grasos; diabetes y acidos grasos, fibra y azucares simples; osteopenia/osteoporosis y calcio y vitamina D; sarcopenia y proteinas, calcio y vitamina D; y entre el deterioro cognitivo y los acidos grasos y los folatos. La muestra 2, asociada a areas rurales, obtuvo puntuaciones en los indices mas bajas. El comportamiento de HADI es similar al de los otros indices (6.24/14 y 6.10/14 en las muestras 1 y 2, respectivamente). Conclusiones: la recopilacion de nutrientes presentada agrega evidencia cientifica util para el diseno de dietas que faciliten un envejecimiento saludable. El nuevo indice propuesto es una herramienta de medicion nutricional especifica en estudios dirigidos a prevenir enfermedades relacionadas con el envejecimiento. EnglishIntroduction: several indexes are used to measure the quality of nutrition in advanced ages. None of them were designed to evaluate nutrition to avoid disabilities in elderly population. Objectives: to retrieve from literature “nutrients and intakes” showing to be involved in aging, and propose a new index, considering this information, to evaluate the quality of nutrition for preventing diseases related to aging. Methods: a bibliographic review was performed, retrieving information on nutrients associated with aging. All these nutrients were incorporated into a new Healthy Aging Diet Index (HADI). Next, a cross-sectional study was carried out with two convenience samples of elderly, collecting the nutritional and dietary data, calculating different validated indexes and comparing them with HADI to validate the results. Results: forty-eight manuscripts were retrieved for full-text analysis. Associations were found between cardiovascular diseases and macronutrients, dietary fibre, sodium and vitamin D; cancer and fatty acids; diabetes and fatty acids, fibre and simple sugars; osteopenia/osteoporosis and calcium and vitamin D; sarcopenia and proteins, calcium, and vitamin D; and between cognitive impairment and fatty acids and folates. Sample 2, associated with rural areas, obtained lower indexes’ scores. The behavior of HADI is similar to the other indexes (6.24/14 and 6.10/14 in samples 1 and 2, respectively). Conclusions: the presented collection of nutrients adds useful evidence for the design of diets that allow healthy aging. The new index proposed is a tool of specific nutritional measurement in studies aimed to prevent diseases related to aging.

5 páginas, 3 figuras, 1 tabla.-- et al., Myopia and hyperopia are at opposite ends of the continuum of refraction, the measure of the eye′s ability to focus light, which is an important cause of visual impairment (when aberrant) and is a highly heritable trait. We conducted a genome-wide association study for refractive error in 4,270 individuals from the TwinsUK cohort. We identified SNPs on 15q25 associated with refractive error (rs8027411, P = 7.91 × 10−8). We replicated this association in six adult cohorts of European ancestry with a combined 13,414 individuals (combined P = 2.07 × 10−9). This locus overlaps the transcription initiation site of RASGRF1, which is highly expressed in neurons and retina and has previously been implicated in retinal function and memory consolidation. Rasgrf1−/− mice show a heavier average crystalline lens (P = 0.001). The identification of a susceptibility locus for refractive error on 15q25 will be important in characterizing the molecular mechanism responsible for the most common cause of visual impairment., The King's College London authors acknowledge funding from the Wellcome Trust, the European Union MyEuropia Marie Curie Research Training Network, Guide Dogs for the Blind Association, the European Community's FP7 (HEALTH-F2-2008-201865-GEFOS), European Network of Genetic and Genomic Epidemiology (ENGAGE) (HEALTH-F4-2007-201413), the FP-5 GenomEUtwin Project (QLG2-CT-2002-01254), US National Institutes of Health (NIH)/National Eye Institute (NEI) grant 1RO1EY018246 and genotyping by the NIH Center for Inherited Disease Research. The study also received support from the National Institute for Health Research (NIHR) comprehensive Biomedical Research Centre award to Guy's and St. Thomas' National Health Service Foundation Trust partnering with King's College London.

The purposes of this review are to study potential biomarkers of exposure for ochratoxin A (OTA) in biological fluids (blood, urine and breast milk) for the period 2005-14, calculate the estimated daily intake (EDI) of OTA by using database consumption for the Spanish population, and, finally, to correlate OTA levels detected in blood and EDI values calculated from food products. The values of OTA detected in potential biomarkers of exposure for blood, breast milk and urine ranged from 0.15 to 18.0, from 0.002 to 13.1, and from 0.013 to 0.2 ng ml(-1), respectively. The calculated EDI for OTA in plasma ranged from 0.15 to 26 ng kg(-1) bw day(-1), higher than that obtained in urine (0.017-0.4 ng kg(-1) bw day(-1)). All these values are correlated with the range of EDI for OTA calculated from food products: 0.0001-25.2 ng kg(-1) bw day(-1).

Melatonin is a hormone mainly involved in the regulation of circadian and seasonal rhythms in both invertebrates and vertebrates. Despite the identification of melatonin in many insects, its involvement in the insect seasonal response remains unclear. A liquid chromatography tandem mass spectrometry (LC-MS/MS) method has been developed for melatonin analysis in aphids (Acyrthosiphon pisum) for the first time. After comparing two different procedures and five extraction solvents, a sample preparation procedure with a mixture of methanol/water (50:50) was selected for melatonin extraction. The method was validated by analyzing melatonin recovery at three spiked concentrations (5, 50 and 100 pg/mg) and showed satisfactory recoveries (75-110%), and good repeatability, expressed as relative standard deviation (

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License ., [Background]: Our prior characterization of RasGrf1 deficient mice uncovered significant defects in pancreatic islet count and size as well as beta cell development and signaling function, raising question about the mechanisms linking RasGrf1 to the generation of those >pancreatic> phenotypes. [Results]: Here, we compared the transcriptional profile of highly purified pancreatic islets from RasGrf1 KO mice to that of WT control animals using commercial oligonucleotide microarrays. RasGrf1 elimination resulted in differential gene expression of numerous components of MAPK- and Calcium-signaling pathways, suggesting a relevant contribution of this GEF to modulation of cellular signaling in the cell lineages integrating the pancreatic islets. Whereas the overall transcriptional profile of pancreatic islets was highly specific in comparison to other organs of the same KO mice, a significant specific repression of Pttg1 was a common transcriptional alteration shared with other tissues of neuroectodermal origin. This observation, together with the remarkable pancreatic phenotypic similarities between RasGrf1 KO and Pttg1 KO mice suggested the possibility of proximal functional regulatory links between RasGrf1 and Pttg1 in pancreatic cell lineages expressing these proteins. [Conclusions]: The specific transcriptional profile and signaling behavior of RasgGrf1 KO pancreatic islets, together with the dominance of Pttg1 over RasGrf1 with regards to the generation of these phenotypes in mouse pancreas, suggest that RasGrf1 is an important upstream component of signal transduction pathways regulating Pttg1 expression and controlling beta cell development and physiological responses., Work supported by grants FIS PI13/02846 and RTICC RD12/0036/0001 from Instituto de Salud Carlos III (ISCIII), and grant SA181U13 from JCyL, Spain. We are grateful to Dr. Douglas Lowy (National Cancer Institute, Bethesda, MD) for providing plasmid pBK-CMV RasGrf1 and to Dr. Shlomo Melmed (Cedars-Sinai Medical Center, Los Angeles, CA) for providing reporter plasmid pGL3-Pttg1 and the single PTTG1 KO mouse strain used in these studies.

Study of dietary supplementation with ENN A mycotoxin during 28 days of exposure time on Wistar rats to determinate its levels in serum, urine and feces and, to evaluate the immunologic effect in peripheral blood lymphocytes (PBL) is presented. The first method for ENN A extraction, determination and detection by LC–MS/MS in serum, urine and feces samples is reported. ENN A food dose administrated was detected in serum samples and influenced lymphocyte phenotyping. Levels in serum were founded from the second week of the experiment; reaching values of 4.76 μg/ml on the fourth week, which corresponds to 3.24 μg/ml in blood. PBL as T helper (CD4 + ) were presented in greater percentages compared to control ( p ≤ 0.001), while T cytotoxic (CD8 + ) decreased significantly compared to control ( p ≤ 0.001). ENN A treatment significantly increased CD4 + /CD3 + and CD4 + /CD8 + ratios but significantly decreased CD8 + /CD3 + ratio. CD4 + /CD8 + ratio was 2.94:1, indicating that PBL surface antigen expression and immune status in Wistar rats treated were impaired by the ENN A mycotoxin.

A 28-day repeated dose preliminary assay, using enniatin A naturally contaminated feed through microbial fermentation by a Fusarium tricinctum strain, was carried out employing two months-old female Wistar rats as in vivo experimental model. In order to simulate a physiological test of a toxic compound naturally produced by fungi, five treated animals were fed during twenty-eight days with fermented feed. As control group, five rats were fed with standard feed. At the 28th day, blood samples were collected for biochemical analysis and the gastrointestinal tract, liver and kidneys were removed from each rat for enniatin A detection and quantitation. Digesta were collected from stomach, duodenum, jejunum, ileum and colon. Enniatin A present in organs and in biological fluids was analyzed by liquid chromatography-diode array detector (LC-DAD) and confirmed by LC-mass spectrometry linear ion trap (MS-LIT); also several serum biochemical parameters and a histological analysis of the duodenal tract were performed. No adverse effects were found in any treated rat at the enniatin A concentration (20.91 mg/kg bw/day) tested during the 28-day experiment. Enniatin A quantitation in biological fluids ranged from 1.50 to 9.00 mg/kg, whereas in the gastrointestinal organs the enniatin A concentration ranged from 2.50 to 23.00 mg/kg. The high enniatin A concentration found in jejunum liquid and tissue points to them as an absorption area. Finally, two enniatin A degradation products were identified in duodenum, jejunum and colon content, probably produced by gut microflora.

This review gives an overview of flow cytometry applications to toxicological studies of several physiological target sites of mycotoxins on different mammalian cell lines. Mycotoxins are secondary metabolites of fungi that may be present in food, feed, air and water. The increasing presence of mycotoxins in crops, their wide distribution in the food chain, and their potential for toxicity demonstrate the need for further knowledge. Flow cytometry has become a valuable tool in mycotoxin studies in recent years for the rapid analysis of single cells in a mixture. In toxicology, the power of these methods lies in the possibility of determining a wide range of cell parameters, providing valuable information to elucidate cell growth and viability, metabolic activity, mitochondrial membrane potential and membrane integrity mechanisms. There are studies using flow cytometry technique on Alternaria, Aspergillus, Fusarium and Penicillium mycotoxins including information about cell type, assay conditions and functional parameters. Most of the studies collected in the literature are on deoxynivalenol and zearalenone mycotoxins. Cell cycle analysis and apoptosis are the processes more widely investigated.