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Here, we present the proof-of-concept of a lateral flow assay (LFA) that is capable of detecting small-molecule targets in a noncompetitive manner by deploying a sandwich-type format based on the aptamer kissing complex (AKC) strategy. A fluorescently labeled hairpin aptamer served as the signaling agent, while a specific RNA hairpin grafted onto the strip served as the capture element. The hairpin aptamer switched from an unfolded to a folded form in the presence of the target, resulting in kissing interactions between the loops of the reporter and the capture agents. This design triggered a target-dependent fluorescent signal at the test line. The AKC-based LFA was developed for the detection of adenosine, achieving a detection limit in the micromolar range. The assay revealed the presence of the same analyte in urine. The method also proved effective with another small molecule (theophylline). We believe that the AKC-based LFA approach could overcome many of the shortcomings associated with conventional signal-off methods and competitive processes.
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Adenosina , Aptámeros de Nucleótidos , Técnicas Biosensibles , Aptámeros de Nucleótidos/química , Adenosina/análisis , Adenosina/orina , Técnicas Biosensibles/métodos , Humanos , Teofilina/análisis , Teofilina/orina , Límite de Detección , Colorantes Fluorescentes/químicaRESUMEN
Glioblastoma (GBM) is the most aggressive malignant glioma, with a very poor prognosis; as such, efforts to explore new treatments and GBM's etiology are a priority. We previously described human GBM cells (R2J-GS) as exhibiting the properties of cancer stem cells (growing in serum-free medium and proliferating into nude mice when orthotopically grafted). Sodium selenite (SS)-an in vitro attractive agent for cancer therapy against GBM-was evaluated in R2J-GS cells. To go further, we launched a preclinical study: SS was given orally, in an escalation-dose study (2.25 to 10.125 mg/kg/day, 5 days on, 2 days off, and 5 days on), to evaluate (1) the absorption of selenium in plasma and organs (brain, kidney, liver, and lung) and (2) the SS toxicity. A 6.75 mg/kg SS dose was chosen to perform a tumor regression assay, followed by MRI, in R2J-GS cells orthotopically implanted in nude mice, as this dose was nontoxic and increased brain selenium concentration. A group receiving TMZ (5 mg/kg) was led in parallel. Although not reaching statistical significance, the group of mice treated with SS showed a slower tumor growth vs. the control group (p = 0.08). No difference was observed between the TMZ and control groups. We provide new insights of the mechanisms of SS and its possible use in chemotherapy.
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Neoplasias Encefálicas/tratamiento farmacológico , Cuerpo Estriado/cirugía , Glioblastoma/tratamiento farmacológico , Células Madre Neoplásicas/trasplante , Selenito de Sodio/efectos adversos , Oligoelementos/efectos adversos , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Animales , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/patología , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cuerpo Estriado/metabolismo , Glioblastoma/patología , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Selenio/metabolismo , Selenito de Sodio/administración & dosificación , Temozolomida/administración & dosificación , Oligoelementos/administración & dosificación , Resultado del TratamientoRESUMEN
We introduced an aptamer switch design that relies on the ability of post-transition/transition metal ions to trigger, through their coordination to nucleobases, substantial DNA destabilization. In the absence of molecular target, the addition of one such metal ion to usual aptamer working solutions promotes the formation of an alternative, inert DNA state. Upon exposure to the cognate compound, the equilibrium is shifted towards the competent DNA form. The switching process was preferentially activated by metal ions of intermediate base over phosphate complexation preference (i.e. Pb2+ , Cd2+ ) and operated with diversely structured DNA molecules. This very simple aptamer switch scheme was applied to the detection of small organics using the fluorescence anisotropy readout mode. We envision that the approach could be adapted to a variety of signalling methods that report on changes in the surface charge density of DNA receptors.
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Many similarities between tryptophan (Trp) and phenylalanine (Phe) metabolisms exist. It is possible that a modification of Trp metabolism might be seen in phenylketonuria (PKU). As some of these metabolites have neuroactive properties, they should be consider in neurological impairment seen in this pathology and not totally explained by blood Phe concentrations. One hundred and fifty-one adult PKU patients (mean age 26.8 years) were included for this study. Plasma Trp, kynurenine (KYN), 3-hydroxykynurenic acid (3HK), and kynurenic acid (KA) were analyzed by liquid chromatography coupled with tandem mass spectrometry. KYN and 3HK were significantly lower in PKU patients compared to general population (P < .0001), and KA was significantly enhanced is this population (P = .009). Furthermore, 3HK concentration was significantly different between PKU patients underwent controlled low-Phe diet compared to PKU patients without this diet (P = .0016). In PKU patients with diet, taking AA substitute enable higher plasma 3HK concentration than without (P = .0008) but still not reaching general population level (P < .0001). Although further study has to be done, it is clear that Trp metabolism is modified in adult PKU patients. An exploration of complete Trp metabolism, and not only Trp concentration, is needed in PKU population, but also in other inborn error of metabolism treated with hypoprotidic diet.
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Fenilcetonurias/sangre , Triptófano/metabolismo , Adulto , Análisis Químico de la Sangre/métodos , Cromatografía Liquida , Femenino , Francia , Humanos , Masculino , Fenilcetonurias/diagnóstico , Fenilcetonurias/metabolismo , Estudios Prospectivos , Espectrometría de Masas en TándemRESUMEN
The aim of this study was to assess the validity of the predictive INTERSALT equation using spot urine samples to estimate 24-h urinary Na (24-hUNa) excretion and daily Na intake among the French adult population. Among 193 French adults ('validation sample'), we assessed the validity by comparing predicted 24-hUNa excretion from spot urine and measured 24-hUNa excretion from 24-h urine collections. Spearman correlation coefficients and Bland-Altman plots were used and we calculated calibration coefficients. In a nationally representative sample of 1720 French adults ('application sample'), the calibrated predictive equation was then applied to the spot urine Na values to estimate 24-hUNa excretion and daily Na intake. In that sample, predicted Na intake was compared with that estimated from 24-h dietary recalls. Results were adjusted and corrected using calibration coefficients. In the validation sample, the measured 24-hUNa excretion was on average 14 % higher than the predicted 24-hUNa (+13 % for men and +16 % for women). Correlation between measured and predicted 24-hUNa excretion was moderate (Spearman r 0·42), and the Bland-Altman plots showed underestimation at lower excretion level and overestimation at higher level. In the application study, estimated daily salt intake was 8·0 g/d using dietary recalls, 8·1 g/d using predicted INTERSALT equation and 9·3 g/d after applying calibration coefficients calculated in the validation study. Despite overall underestimation of 24-hUNa excretion by spot urinary Na, the use of predictive INTERSALT equation remains an acceptable alternative in monitoring global Na intake/excreted in the French population but its use is not advised at the individual level.
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Sodio en la Dieta/administración & dosificación , Sodio/orina , Adulto , Anciano , Dieta , Registros de Dieta , Reacciones Falso Negativas , Femenino , Francia , Humanos , Masculino , Recuerdo Mental , Persona de Mediana Edad , Encuestas Nutricionales , Factores de Tiempo , Toma de Muestras de Orina/métodosRESUMEN
Obstructive sleep apnoea syndrome is characterized by repetitive episodes of upper airway collapse during sleep resulting in chronic intermittent hypoxia (IH). Obstructive sleep apnoea syndrome, through IH, promotes cardiovascular and metabolic disorders. Endothelin-1 (ET-1) secretion is upregulated by IH, and is able to modulate adipocyte metabolism. Therefore, the present study aimed to characterize the role of ET-1 in the metabolic consequences of IH on adipose tissue in vivo and in vitro. Wistar rats were submitted to 14 days of IH-cycles (30 s of 21% FiO2 and 30 s of 5% FiO2 ; 8 h day(-1) ) or normoxia (air-air cycles) and were treated or not with bosentan, a dual type A and B endothelin receptor (ETA-R and ETB-R) antagonist. Bosentan treatment decreased plasma free fatty acid and triglyceride levels, and inhibited IH-induced lipolysis in adipose tissue. Moreover, IH induced a 2-fold increase in ET-1 transcription and ETA-R expression in adipose tissue that was reversed by bosentan. In 3T3-L1 adipocytes, ET-1 upregulated its own and its ETA-R transcription and this effect was abolished by bosentan. Moreover, ET-1 induced glycerol release and inhibited insulin-induced glucose uptake. Bosentan and BQ123 inhibited these effects. Bosentan also reversed the ET-1-induced phosphorylation of hormone-sensitive lipase (HSL) on Ser(660) . Finally, ET-1-induced lipolysis and HSL phosphorylation were also observed under hypoxia. Altogether, these data suggest that ET-1 is involved in IH-induced lipolysis in Wistar rats, and that upregulation of ET-1 production and ETA-R expression by ET-1 itself under IH could amplify its effects. Moreover, ET-1-induced lipolysis could be mediated through ETA-R and activation of HSL by Ser(660) phosphorylation.
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Tejido Adiposo/metabolismo , Endotelina-1/metabolismo , Hipoxia/metabolismo , Lipólisis , Procesamiento Proteico-Postraduccional , Esterol Esterasa/metabolismo , Células 3T3 , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/patología , Animales , Bosentán , Antagonistas de los Receptores de Endotelina/farmacología , Endotelina-1/genética , Ácidos Grasos/sangre , Hipoxia/patología , Masculino , Ratones , Fosforilación , Ratas , Ratas Wistar , Receptores de Endotelina/genética , Receptores de Endotelina/metabolismo , Sulfonamidas/farmacología , Triglicéridos/sangreRESUMEN
We report herein a novel sandwich-type enzyme-linked assay for the "signal-on" colorimetric detection of small molecules. The approach (referred to as enzyme-linked aptamer kissing complex assay (ELAKCA)) relied on the kissing complex-based recognition of the target-bound hairpin aptamer conformational state by a specific RNA hairpin probe. The aptamer was covalently immobilized on a microplate well surface to act as target capture element. Upon small analyte addition, the folded aptamer was able to bind to the biotinylated RNA hairpin module through loop-loop interaction. The formed ternary complex was then revealed by the introduction of the streptavidin-horseradish peroxidase conjugate that catalytically converted the 3,3',5,5'-tetramethylbenzidine substrate into a colorimetric product. ELAKCA was successfully designed for two different systems allowing detecting the adenosine and theophylline molecules. The potential practical applicability in terms of biological sample analysis (human plasma), temporal stability, and reusability was also reported. Owing to the variety of both hairpin functional nucleic acids, kissing motifs, and enzyme-based signaling systems, ELAKCA opens up new prospects for developing small molecule sensing platforms of wide applications.
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Pruebas de Enzimas/métodos , Adenosina/sangre , Aptámeros de Nucleótidos/química , Bencidinas/química , Compuestos Cromogénicos/química , Colorimetría , Pruebas de Enzimas/instrumentación , Peroxidasa de Rábano Silvestre/química , Humanos , Estreptavidina/química , Teofilina/sangreRESUMEN
NEW FINDINGS: What is the central question of this study? This study addresses the relative impact of obesity and intermittent hypoxia in the pathophysiological process of obstructive sleep apnoea by investigating the metabolic, inflammatory and cardiovascular consequences of intermittent hypoxia in lean and obese Zucker rats. What is the main finding and its importance? We found that obesity and intermittent hypoxia have mainly distinct consequences on the investigated inflammatory and cardiometabolic parameters in Zucker rats. This suggests that, for a given severity of sleep apnea, the association of obesity and obstructive sleep apnoea may not necessarily be deleterious. Obstructive sleep apnoea is associated with obesity with a high prevalence, and both co-morbidities are independent cardiovascular risk factors. Intermittent hypoxia (IH) is thought to be the main factor responsible for the obstructive sleep apnoea-related cardiometabolic alterations. The aim of this study was to assess the respective impact of obesity and IH on the inflammatory and cardiometabolic state in rats. Lean and obese Zucker rats were exposed to normoxia or chronic IH, and we assessed metabolic and inflammatory parameters, such as plasma lipids and glucose, serum leptin and adiponectin, liver cytokines, nuclear factor-κB activity and cardiac endothelin-1 levels. Myocardial infarct size was also evaluated following in vitro ischaemia-reperfusion. Circulating lipids, insulin, homeostasis model assessment of insulin resistance (HOMA-IR), leptin and adiponectin levels were higher in obese versus lean rats. Chronic IH did not have a significant impact on metabolic parameters in lean rats. In obese rats, IH increased glycaemia and HOMA-IR. Liver interleukin-6 and tumour necrosis factor-α levels were elevated in lean rats exposed to IH; obesity prevented the increase in interleukin-6 but not in tumour necrosis factor-α. Finally, IH exposure enhanced myocardial sensitivity to infarction in both lean and obese rats and increased cardiac endothelin-1 in lean but not obese rats. In conclusion, this study shows that the dyslipidaemia and insulin resistance induced by obesity of genetic origin does not enhance the deleterious cardiovascular response to IH and may even partly protect against IH-induced inflammation.
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Enfermedades Cardiovasculares/metabolismo , Hipoxia/metabolismo , Inflamación/metabolismo , Obesidad/metabolismo , Adiponectina/metabolismo , Animales , Glucemia/metabolismo , Enfermedades Cardiovasculares/fisiopatología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Endotelina-1/metabolismo , Insulina/metabolismo , Interleucina-6/metabolismo , Leptina/sangre , Lípidos/sangre , Hígado/metabolismo , Masculino , Miocardio/metabolismo , FN-kappa B/metabolismo , Ratas , Ratas Zucker , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
BACKGROUND: Pomegranate peel extract (PPE) contains several compounds with antioxidative properties. PPE added to foods may interact with endogenous antioxidants and promote health. However, little is known about the biochemical mechanisms by which PPE exerts their actions on tissues of biological systems in vivo. The purpose of this study was to determine the effects of PPE on activities of antioxidant enzymes. Mice were used to investigate the effects of PPE on plasma levels of malondialdehyde (MDA), tissue MDA content and activities of superoxide dismutase 1 (SOD1), SOD2 and glutathione peroxidase (GPX) in the small intestine, liver and skeletal muscle - different tissues involved in the digestion, absorption and metabolism of dietary nutrients. Control mice were fed a standard diet, whereas treated mice were fed for 40 days with the standard diet containing 5% or 10% PPE. RESULTS: Mice fed the 10% PPE diet exhibited lower plasma MDA concentrations, reduced content of MDA in the small intestine and liver and higher levels of SOD1 and GPX activities in the small intestine compared to mice fed the control diet. CONCLUSIONS: These findings demonstrate that intake of PPE in diet attenuates small intestine lipid peroxidation and strengthens the first line of small intestine antioxidant defense by enhancing enzymatic antioxidative pathways. PPE is worthy of further study as a therapeutic approach to prevent peroxidative stress-induced gut pathogenesis. © 2015 Society of Chemical Industry.
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Antioxidantes/administración & dosificación , Intestino Delgado/metabolismo , Peroxidación de Lípido/efectos de los fármacos , Lythraceae/química , Extractos Vegetales/farmacología , Animales , Antioxidantes/farmacología , Dieta , Femenino , Frutas , Glutatión Peroxidasa/metabolismo , Intestino Delgado/enzimología , Hígado/metabolismo , Malondialdehído/sangre , Ratones , Músculo Esquelético/metabolismo , Extractos Vegetales/administración & dosificación , Superóxido Dismutasa/metabolismoRESUMEN
New technologies are promising for the use of short-term instruments for dietary data collection; however, innovative tools should be validated against objective biomarkers. The aim of the present study was to investigate the validity of a Web-based, self-administered dietary record (DR) tool using protein, K and Na intakes against 24 h urinary biomarkers (24 h U). Of the total participants, 199 adult volunteers (104 men and 95 women, mean age 50·5 (23-83 years)) of the NutriNet-Santé Study were included in the protocol. They completed three non-consecutive-day DR and two 24 h U on the first and third DR days. Relative differences between reported (DR) and measured (24 h U) intakes were calculated from the log ratio (DR/24 h U) for protein, K and Na intakes: -14·4,+2·6 and -2·1 % for men; and -13·9, -3·7 and -8·3 % for women, respectively. The correlations between reported and true intakes were 0·61, 0·78 and 0·47 for men and 0·64, 0·42 and 0·37 for women for protein, K and Na, respectively. Attenuation factors, that represent attenuation of the true diet-disease relationship due to measurement error (a value closer to 1 indicating lower attenuation), ranged from 0·23 (Na, women) to 0·60 (K, men). We showed that the Web-based DR tool used in the NutriNet-Santé cohort study performs well in estimating protein and K intakes and fairly well in estimating Na intake. Furthermore, three non-consecutive-day DR appear to be valid for estimating usual intakes of protein and K, although caution is advised regarding the generalisability of these findings to other nutrients and general population.
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Dieta , Proteínas en la Dieta/administración & dosificación , Potasio en la Dieta/administración & dosificación , Sodio en la Dieta/administración & dosificación , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/orina , Estudios de Cohortes , Dieta/efectos adversos , Registros de Dieta , Femenino , Humanos , Internet , Masculino , Persona de Mediana Edad , Nitrógeno/orina , Evaluación Nutricional , Paris , Potasio/orina , Autoinforme , Sodio/orina , Adulto JovenRESUMEN
In this paper, we introduce a geometric method for 3D reconstruction of the exterior environment using a panoramic microwave radar and a camera. We rely on the complementarity of these two sensors considering the robustness to the environmental conditions and depth detection ability of the radar, on the one hand, and the high spatial resolution of a vision sensor, on the other. Firstly, geometric modeling of each sensor and of the entire system is presented. Secondly, we address the global calibration problem, which consists of finding the exact transformation between the sensors' coordinate systems. Two implementation methods are proposed and compared, based on the optimization of a non-linear criterion obtained from a set of radar-to-image target correspondences. Unlike existing methods, no special configuration of the 3D points is required for calibration. This makes the methods flexible and easy to use by a non-expert operator. Finally, we present a very simple, yet robust 3D reconstruction method based on the sensors' geometry. This method enables one to reconstruct observed features in 3D using one acquisition (static sensor), which is not always met in the state of the art for outdoor scene reconstruction. The proposed methods have been validated with synthetic and real data.
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BACKGROUND & AIMS: Fibrosis blood tests have been validated in chronic hepatitis C. Their diagnostic accuracy is less documented in hepatitis B. The aim of this study was to describe the diagnostic performance of FibroTest®, FibroMeter®, and HepaScore® for liver fibrosis in hepatitis B compared to hepatitis C. METHODS: 510 patients mono-infected with hepatitis B or C and matched on fibrosis stage were included. Blood tests were performed the day of the liver biopsy. Histological lesions were staged according to METAVIR. RESULTS: Fibrosis stages were distributed as followed: F0 n=76, F1 n=192, F2 n=132, F3 n=54, F4 n=56. Overall diagnostic performance of blood tests were similar between hepatitis B and C with AUROC ranging from 0.75 to 0.84 for significant fibrosis, 0.82 to 0.85 for extensive fibrosis and 0.84 to 0.87 for cirrhosis. Optimal cut-offs were consistently lower in hepatitis B compared to hepatitis C, especially for the diagnosis of extensive fibrosis and cirrhosis, with decreased sensitivity and negative predictive values. More hepatitis B than C patients with F ⩾3 were underestimated: FibroTest®: 47% vs. 26%, FibroMeter®: 24% vs. 6%, HepaScore®: 41% vs. 24%, p<0.01. Multivariate analysis showed that hepatitis B (0R 3.4, 95% CI 1.2-19.2, p<0.02) and low γGT (OR 7.3, 95% CI 2.0-27.0, p<0.003) were associated with fibrosis underestimation. CONCLUSION: Overall the diagnostic performance of blood tests is similar in hepatitis B and C. The risk of underestimating significant fibrosis and cirrhosis is however greater in hepatitis B and cannot be entirely corrected by the use of more stringent cut-offs.
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Hepatitis B Crónica/sangre , Hepatitis B Crónica/patología , Hepatitis C Crónica/sangre , Hepatitis C Crónica/patología , Cirrosis Hepática/sangre , Cirrosis Hepática/patología , Adulto , Alanina Transaminasa/sangre , Femenino , Pruebas Hematológicas/métodos , Humanos , Hígado/patología , Masculino , Persona de Mediana Edad , Estudios Prospectivos , gamma-Glutamiltransferasa/sangreAsunto(s)
Leucemia Mieloide Aguda/sangre , Leucemia Mieloide Aguda/genética , Oxidación-Reducción , Adulto , Anciano , Antimicina A/farmacología , Antioxidantes/metabolismo , Femenino , Humanos , Estimación de Kaplan-Meier , Leucemia Mieloide Aguda/mortalidad , Peroxidación de Lípido , Masculino , Persona de Mediana Edad , Mitocondrias/metabolismo , NADPH Oxidasas/metabolismo , Compuestos Onio/farmacología , Pronóstico , Modelos de Riesgos Proporcionales , Estudios Prospectivos , Especies Reactivas de Oxígeno/metabolismo , Rotenona/farmacología , Acetato de Tetradecanoilforbol/farmacología , Translocación GenéticaRESUMEN
BACKGROUND/AIMS: Low-grade inflammation is an independent risk factor for cardiovascular disease. Relationships between the antioxidant status and inflammatory biomarkers could give new insights into cardiovascular disease prevention. We investigated long-term associations between the antioxidant nutrient (vitamin C, α-tocopherol, ß-carotene) status and C-reactive protein (CRP) in a population-based cohort. METHODS: Subjects included in the French SU.VI.MAX trial study who had available data on baseline (1994-1995) blood nutrient concentrations and CRP measurements 12 years later (2007-2009) were included. Associations between baseline antioxidant circulating concentrations and elevated CRP (>3 mg/l) were investigated in multivariate logistic regression models. Subgroup analyses were performed according to gender, supplementation group of the initial trial, smoking status, and alcohol intake. RESULTS: Serum α-tocopherol (n = 2,060) and vitamin C (n = 1,719) concentrations [odds ratio (OR) and 95% confidence interval (95% CI) quintile 5 vs. 1: OR 1.10 (95% CI 0.71-1.73), p for trend = 0.533, vs. OR 0.79 (95% CI 0.48-1.29), p for trend = 0.121, respectively] were not associated with elevated CRP concentrations. The ß-carotene status (n = 2,048) was inversely associated with elevated CRP [adjusted OR quintile 5 vs. 1: OR 0.61 (95% CI 0.38-0.98), p for trend = 0.01]. Subgroup analyses showed that associations were stronger in women (p for trend = 0.004), never smokers (p for trend = 0.009) and subjects in the supplementation group (p for trend = 0.002). CONCLUSIONS: Our results suggest that the ß-carotene status may be inversely associated with low-grade inflammation in the long term.
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Antioxidantes/metabolismo , Proteína C-Reactiva/metabolismo , Inflamación/epidemiología , Ácido Ascórbico/sangre , Estudios de Cohortes , Femenino , Francia/epidemiología , Humanos , Inflamación/sangre , Masculino , Persona de Mediana Edad , Ensayos Clínicos Controlados Aleatorios como Asunto , Factores de Riesgo , alfa-Tocoferol/sangre , beta Caroteno/sangreRESUMEN
Iron is an essential element to the well functionning of the organism and requires careful maintenance of its homeostasis. This is mainly due to hepcidin, a hormone secreted by the liver that controls the flow of iron within the body. It has a hyposideremic action by reducing the expression of ferroportin, the only protein known to this day, which can export iron into the extracellular environment. This has the effect of decreasing intestinal absorption and increasing intracellular retention, especially in macrophages. Hepcidin is stimulated by elevation of iron and inflammation while activation of erythropoiesis inhibits it. Understanding its regulation allows a better understanding of the pathophysiological mechanisms of overload diseases and iron deficiency. Therefore, hepcidin analysis is interesting for the exploration of iron homeostasis. In combination with other biological parameters of iron status, it is possible to find better instructions for therapeutic managements of iron metabolism diseases. Thus, an assaying method by coupling liquid chromatography and tandem mass spectrometry has been implemented at Grenoble University Hospital and the analytical performances of this assay met the laboratory requirements in terms of reliability of the analysis.
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Hepcidinas , Hierro , Humanos , Hepcidinas/metabolismo , Homeostasis/fisiología , Hígado/metabolismo , Reproducibilidad de los ResultadosRESUMEN
Herein, we report a novel approach for the design of a colorimetric aptasensor, relying on a Dye Salt Aggregation-based Colorimetric Oligonucleotide assay (DYSACO assay). This method is based on the use of an intercalating agent, Nile Blue (NB), whose aggregation capacities (and thus modification of its absorption spectrum) are drastically amplified by adding salts to the working solution. The presence of an aptamer could protect NB from such aggregation process due to its intercalation into double-stranded DNA and/or interaction with nucleobases. In response to the addition of the specific ligand, the competition between NB and the target for binding to the aptamer occurs, resulting in an increase in the dye salt aggregation and then in the blue-to-blank color change of the solution. The proof-of-principle was demonstrated by employing the anti-l-tyrosinamide aptamer and the assay was successfully applied to the trace enantiomer detection, allowing the detection of an enantiomeric impurity down to approximately 2% in a non-racemic sample. Through a reversed mechanism based on the increased capture of NB by DNA upon analyte binding, the sensing platform was further demonstrated for the Hg(II) detection. Water samples of different origin were spiked with Hg(II) analyte at final range concentrations comprised between (0.5-15 µM). An excellent overall recovery of 122 ± 14%; 105 ± 14%; 99 ± 9%; was respectively obtained from river, tap and mineral water, suggesting that the sensor can be used under real sample conditions. The assay was also shown to work for sensing the ochratoxin A and d-arginine vasopressin compounds, revealing its simplicity and generalizability potentialities.
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Aptámeros de Nucleótidos , Técnicas Biosensibles , Mercurio , Nanopartículas del Metal , Colorimetría/métodos , Nanopartículas del Metal/química , Técnicas Biosensibles/métodos , Oro/química , Cloruro de Sodio , ADN/química , Péptidos , Cloruro de Sodio Dietético , Aptámeros de Nucleótidos/químicaRESUMEN
BACKGROUND: In vitro and toxicological studies have shown that non-persistent environmental chemicals can perturb thyroid hormone homeostasis. Epidemiological studies with improved exposure assessment (i.e., repeated urine samples) are needed to evaluate effects of these compounds, individually or as a mixture, in humans. We studied the associations between prenatal exposure to non-persistent environmental chemicals and neonatal thyroid hormones. METHODS: The study population consisted of 442 mother-child pairs from the French SEPAGES mother-child cohort recruited between July 2014 and July 2017. For each participant, four parabens, five bisphenols, triclosan, triclocarban, benzophenone-3 as well as metabolites of phthalates and of di(isononyl)cyclohexane-1,2-dicarboxylate were assessed in two pools of repeated urine samples (median: 21 spot urines per pool), collected in the 2nd and 3rd trimesters of pregnancy, respectively. Thyroid stimulating hormone (TSH) and total thyroxine (T4) levels were determined in newborns from a heel-prick blood spot. Maternal iodine and selenium were assessed in urine and serum, respectively. Adjusted linear regression (uni-pollutant model) and Bayesian Kernel Machine Regression (BKMR, mixture model) were applied to study overall and sex-stratified associations between chemicals and hormone concentrations. RESULTS: Interaction with child sex was detected for several compounds. Triclosan, three parabens, and one phthalate metabolite (OH-MPHP) were negatively associated with T4 among girls in the uni-pollutant model. BKMR also suggested a negative association between the mixture and T4 in girls, whereas in boys the association was positive. The mixture was not linked to TSH levels, and for this hormone the uni-pollutant model revealed associations with only a few compounds. CONCLUSION: Our study, based on repeated urine samples to assess exposure, showed that prenatal exposure to some phenols and phthalates disturb thyroid hormone homeostasis at birth. Furthermore, both uni-pollutant and mixture models, suggested effect modification by child sex, while, to date underlying mechanisms for such sex-differences are not well understood.