Electrochemical As(III) whole-cell based biochip sensor.

The development of a whole-cell based sensor for arsenite detection coupling biological engineering and electrochemical techniques is presented. This strategy takes advantage of the natural Escherichia coli resistance mechanism against toxic arsenic species, such as arsenite, which consists of the selective intracellular recognition of arsenite and its pumping out from the cell. A whole-cell based biosensor can be produced by coupling the intracellular recognition of arsenite to the generation of an electrochemical signal. Hereto, E. coli was equipped with a genetic circuit in which synthesis of beta-galactosidase is under control of the arsenite-derepressable arsR-promoter. The E. coli reporter strain was filled in a microchip containing 16 independent electrochemical cells (i.e. two-electrode cell), which was then employed for analysis of tap and groundwater samples. The developed arsenic-sensitive electrochemical biochip is easy to use and outperforms state-of-the-art bacterial bioreporters assays specifically in its simplicity and response time, while keeping a very good limit of detection in tap water, i.e. 0.8ppb. Additionally, a very good linear response in the ranges of concentration tested (0.94ppb to 3.75ppb, R(2)=0.9975 and 3.75 ppb to 30ppb, R(2)=0.9991) was obtained, complying perfectly with the acceptable arsenic concentration limits defined by the World Health Organization for drinking water samples (i.e. 10ppb). Therefore, the proposed assay provides a very good alternative for the portable quantification of As (III) in water as corroborated by the analysis of natural groundwater samples from Swiss mountains, which showed a very good agreement with the results obtained by atomic absorption spectroscopy.

Procalcitonin and C-reactive protein in pericardial fluid for postmortem diagnosis of sepsis.

The aim of this study was to investigate the presence and concentrations of procalcitonin and C-reactive protein in pericardial fluid and compare these levels to those found in the postmortem serum obtained from the femoral blood. Two groups were formed, a sepsis-related fatalities group and a control group. Postmortem native CT scans, autopsies, histology, neuropathology and toxicology as well as other postmortem biochemistry investigations were performed in all cases. Pericardial fluid procalcitonin levels were significantly different between the cases of sepsis-related fatalities and those of the control group. Postmortem serum procalcitonin levels below the detection limit were also reflected in undetectable pericardial fluid levels. Similarly, a large increase in postmortem serum procalcitonin levels was reflected in a large increase of procalcitonin pericardial fluid levels. Based on these findings, pericardial fluid could be an alternative to postmortem serum for the determination of procalcitonin levels in cases where postmortem serum is not available and measurements of procalcitonin are required to circumstantiate the pathogenesis of death.

Assessment of driving capability through the use of clinical and psychomotor tests in relation to blood cannabinoids levels following oral administration of 20 mg dronabinol or of a cannabis decoction made with 20 or 60 mg Delta9-THC.

Delta(9)-Tetrahydrocannabinol (THC) is frequently found in the blood of drivers suspected of driving under the influence of cannabis or involved in traffic crashes. The present study used a double-blind crossover design to compare the effects of medium (16.5 mg THC) and high doses (45.7 mg THC) of hemp milk decoctions or of a medium dose of dronabinol (20 mg synthetic THC, Marinol on several skills required for safe driving. Forensic interpretation of cannabinoids blood concentrations were attempted using the models proposed by Daldrup (cannabis influencing factor or CIF) and Huestis and coworkers. First, the time concentration-profiles of THC, 11-hydroxy-Delta(9)-tetrahydrocannabinol (11-OH-THC) (active metabolite of THC), and 11-nor-9-carboxy-Delta(9)-tetrahydrocannabinol (THCCOOH) in whole blood were determined by gas chromatography-mass spectrometry-negative ion chemical ionization. Compared to smoking studies, relatively low concentrations were measured in blood. The highest mean THC concentration (8.4 ng/mL) was achieved 1 h after ingestion of the strongest decoction. Mean maximum 11-OH-THC level (12.3 ng/mL) slightly exceeded that of THC. THCCOOH reached its highest mean concentration (66.2 ng/mL) 2.5-5.5 h after intake. Individual blood levels showed considerable intersubject variability. The willingness to drive was influenced by the importance of the requested task. Under significant cannabinoids influence, the participants refused to drive when they were asked whether they would agree to accomplish several unimportant tasks, (e.g., driving a friend to a party). Most of the participants reported a significant feeling of intoxication and did not appreciate the effects, notably those felt after drinking the strongest decoction. Road sign and tracking testing revealed obvious and statistically significant differences between placebo and treatments. A marked impairment was detected after ingestion of the strongest decoction. A CIF value, which relies on the molar ratio of main active to inactive cannabinoids, greater than 10 was found to correlate with a strong feeling of intoxication. It also matched with a significant decrease in the willingness to drive, and it matched also with a significant impairment in tracking performances. The mathematic model II proposed by Huestis et al. (1992) provided at best a rough estimate of the time of oral administration with 27% of actual values being out of range of the 95% confidence interval. The sum of THC and 11-OH-THC blood concentrations provided a better estimate of impairment than THC alone. This controlled clinical study points out the negative influence on fitness to drive after medium or high dose oral THC or dronabinol.

Antihypertensive drug treatment changes in the general population: the CoLaus study.

BACKGROUND: Changes in antihypertensive drug treatment are paramount in the adequate management of patients with hypertension, still, there is little information regarding changes in antihypertensive drug treatment in Switzerland. Our aim was to assess those changes and associated factors in a population-based, prospective study. METHODS: Data from the population-based, CoLaus study, conducted among subjects initially aged 35-75 years and living in Lausanne, Switzerland. 772 hypertensive subjects (371 women) were followed for a median of 5.4 years. Data Subjects were defined as continuers (no change), switchers (one antihypertensive class replaced by another), combiners (one antihypertensive class added) and discontinuers (stopped treatment). The distribution and the factors associated with changes in antihypertensive drug treatment were assessed. RESULTS: During the study period, the prescription of diuretics decreased and of ARBs increased: at baseline, diuretics were taken by 46.9% of patients; angiotensin receptor blockers (ARB) by 44.7%, angiotensin converting enzyme inhibitors (ACEI) by 28.8%, beta-blockers (BB) by 28.0%, calcium channel blockers (CCB) by 18.9% and other antihypertensive drugs by 0.3%. At follow-up (approximately 5 years later), their corresponding percentages were 42.8%, 51.7%, 25.5%, 33.0% 20.7% and 1.0%. Among all participants, 54.4% (95% confidence interval: 50.8-58.0) were continuers, 26.9% (23.8-30.2) combiners, 12.7% (10.4-15.3) switchers and 6.0% (4.4-7.9) discontinuers. Combiners had higher systolic blood pressure values at baseline than the other groups (p < 0.05). Almost one third (30.6%) of switchers and 29.3% of combiners improved their blood pressure status at follow-up, versus 18.8% of continuers and 8.7% of discontinuers (p < 0.001). Conversely, almost one third (28.3%) of discontinuers became hypertensive (systolic ≥140 mm Hg or diastolic ≥90 mm Hg), vs. 22.1% of continuers, 16.3% of switchers and 11.5% of combiners (p < 0.001). Multivariate analysis showed baseline uncontrolled hypertension, ARBs, drug regimen (monotherapy/polytherapy) and overweight/obesity to be associated with changes in antihypertensive therapy. CONCLUSION: In Switzerland, ARBs have replaced diuretics as the most commonly prescribed antihypertensive drug. Uncontrolled hypertension, ARBs, drug regimen (monotherapy or polytherapy) and overweight/obesity are associated with changes in antihypertensive treatment.

Sex differences in urinary levels of several biological indicators of exposure: a human volunteer study

The aim of the study was to quantify the variability on biological indicators of exposure between men and women for three well known solvents: methyl ethyl ketone, 1-methoxy-2-propanol and 1,1,1-trichloroethane. Another purpose was to explore the effect of selected CYP2E1 polymorphisms on the toxicokinetic profile. Controlled human exposures were carried out in a 12m(3) exposure chamber for each solvent separately, during 6h and at half of the threshold limit value. The human volunteers groups were composed of ten young men and fifteen young women, including ten women using hormonal contraceptive. An analysis of variance mainly showed an effect on the urinary levels of several biomarkers of exposure among women due to the use of hormonal contraceptive, with an increase of more than 50% in metabolites concentrations and a decrease of up to 50% in unchanged substances concentrations, suggesting an increase in their metabolism rate. The results also showed a difference due to the genotype CYP2E1*6, when exposed to methyl ethyl ketone, with a tendency to increase CYP2E1 activity when volunteers were carriers of the mutant allele. Our study suggests that not only physiological differences between men and women but also differences due to sex hormones levels can have an impact on urinary concentrations of several biomarkers of exposure. The observed variability due to sex among biological exposure indices can lead to misinterpretation of biomonitoring results. This aspect should have its place in the approaches for setting limits of occupational exposure. [Authors]

Immediate and delayed effects of subchronic Paraquat exposure during an early differentiation stage in 3D-rat brain cell cultures.

Xenobiotic exposure is a risk factor in the etiology of neurodegenerative disease. It was recently hypothesized that restricted exposure during brain development could predispose for a neurodegenerative disease later in life. As neuroinflammation contributes to progressive neurodegeneration, it is suspected that neurodevelopmental xenobiotic exposure could elicit a neuroinflammatory process, which over time may assume a detrimental character. We investigated the neurotoxic effects of paraquat (PQ) in three-dimensional whole rat brain cell cultures, exposed during an early differentiation stage, comparing immediate effects-directly post exposure-with long-term effects, 20 days after interrupted PQ-administration. Adverse effects and neuroinflammatory responses were assessed by measuring changes in gene- and protein-expression as well as by determining cell morphology changes. Differentiating neural cultures were highly susceptible to PQ and showed neuronal damage and strong astrogliosis. After the 20-day washout period, neurons partially recovered, whereas astrogliosis persisted, and was accompanied by microglial activation of a neurodegenerative phenotype. Our data shows that immediate and long-term effects of subchronic PQ-exposure differ. Also, PQ-exposure during this window of extensive neuronal differentiation led to a delayed microglial activation, of a character that could promote further pro-inflammatory signals that enable prolonged inflammation, thereby fueling further neurodegeneration.

The permeability P-glycoprotein: a focus on enantioselectivity and brain distribution.

IMPORTANCE OF THE FIELD: The permeability glycoprotein (P-gp) is an important protein transporter involved in the disposition of many drugs with different chemical structures, but few studies have examined a possible stereoselectivity in its activity. P-gp can have a major impact on the distribution of drugs in selected organs, including the brain. Polymorphisms of the ABCB1 gene, which encodes for P-gp, can influence the kinetics of several drugs. AREAS COVERED IN THIS REVIEW: A search including publications from 1990 up to 2009 was performed on P-gp stereoselectivity and on the impact of ABCB1 polymorphisms on enantiomer brain distribution. WHAT THE READER WILL GAIN: Despite stereoselectivity not being expected because of the large variability of chemical structures of P-gp substrates, structure-activity relationships suggest different P-gp-binding sites for enantiomers. Enantioselectivity in the activity of P-gp has been demonstrated by in vitro studies and in animal models (preferential transport of one enantiomer or different inhibitory potencies towards P-gp activity between enantiomers). There is also in vivo evidence of an enantioselective drug transport at the human blood-brain barrier. TAKE HOME MESSAGE: The significant enantioselective activity of P-gp might be clinically relevant and must be taken into account in future studies.

Characterization and classification of matrix effects in biological samples analyses.

An exhaustive classification of matrix effects occurring when a sample preparation is performed prior to liquid-chromatography coupled to mass spectrometry (LC-MS) analyses was proposed. A total of eight different situations were identified allowing the recognition of the matrix effect typology via the calculation of four recovery values. A set of 198 compounds was used to evaluate matrix effects after solid phase extraction (SPE) from plasma or urine samples prior to LC-ESI-MS analysis. Matrix effect identification was achieved for all compounds and classified through an organization chart. Only 17% of the tested compounds did not present significant matrix effects.

Toxic substances in blood: an analysis of current recommendations under a Bayesian (decision) approach

This article extends existing discussion in literature on probabilistic inference and decision making with respect to continuous hypotheses that are prevalent in forensic toxicology. As a main aim, this research investigates the properties of a widely followed approach for quantifying the level of toxic substances in blood samples, and to compare this procedure with a Bayesian probabilistic approach. As an example, attention is confined to the presence of toxic substances, such as THC, in blood from car drivers. In this context, the interpretation of results from laboratory analyses needs to take into account legal requirements for establishing the 'presence' of target substances in blood. In a first part, the performance of the proposed Bayesian model for the estimation of an unknown parameter (here, the amount of a toxic substance) is illustrated and compared with the currently used method. The model is then used in a second part to approach-in a rational way-the decision component of the problem, that is judicial questions of the kind 'Is the quantity of THC measured in the blood over the legal threshold of 1.5 μg/l?'. This is pointed out through a practical example.

Biomarkers of oxidative stress and its association with the urinary reducing capacity in bus maintenance workers.

BACKGROUND: Exposure to particles (PM) induces adverse health effects (cancer, cardiovascular and pulmonary diseases). A key-role in these adverse effects seems to be played by oxidative stress, which is an excess of reactive oxygen species relative to the amount of reducing species (including antioxidants), the first line of defense against reactive oxygen species. The aim of this study was to document the oxidative stress caused by exposure to respirable particles in vivo, and to test whether exposed workers presented changes in their urinary levels for reducing species.METHODS: Bus depot workers (n = 32) exposed to particles and pollutants (respirable PM4, organic and elemental carbon, particulate metal content, polycyclic aromatic hydrocarbons, NOx, O3) were surveyed over two consecutive days. We collected urine samples before and after each shift, and quantified an oxidative stress biomarker (8-hydroxy-2'-deoxyguanosine), the reducing capacity and a biomarker of PAH exposure (1-hydroxypyrene). We used a linear mixed model to test for associations between the oxidative stress status of the workers and their particle exposure as well as with their urinary level of reducing species.RESULTS: Workers were exposed to low levels of respirable PM4 (range 25-71 μg/m3). However, urinary levels of 8-hydroxy-2'-deoxyguanosine increased significantly within each shift and between both days for non-smokers. The between-day increase was significantly correlated (p < 0.001) with the concentrations of organic carbon, NOx, and the particulate copper content. The within-shift increase in 8OHdG was highly correlated to an increase of the urinary reducing capacity (Spearman ρ = 0.59, p < 0.0001).CONCLUSIONS: These findings confirm that exposure to components associated to respirable particulate matter causes a systemic oxidative stress, as measured with the urinary 8OHdG. The strong association observed between urinary 8OHdG with the reducing capacity is suggestive of protective or other mechanisms, including circadian effects. Additional investigations should be performed to understand these observations.

Evaluation of the toxicity of different metal compounds in the developing brain using aggregating cell cultures as a model.

To evaluate their toxicity in the developing brain, eight metal compounds, [bismuth sodium tartrate (BiNA-tartrate), CdCl(2), CoCl(2), HgCl(2), dimethyl mercury, NiCl(2), TlCl and triethyltin chloride (TET)] were tested in aggregating cell cultures of foetal rat telencephalon. The compounds were applied to the cultures continuously, either during an early developmental stage (between days 5 and 14) or during and advanced stage of maturation (between days 24 and 34). Changes in the activities of cell type-specific enzymes were used as a criterion for toxicity. A general cytotoxic effect was observed after treatment with either CdCl(2), HgCl(2) or TET at 10(-6)m, and with TlCl at 10(-5)m. Selective effects were found with BiNa-tartrate and dimethylmercury. CoCl(2) did not modify the parameters tested, whereas a stimulant effect was found with NiCl(2). The effects of several compounds were development dependent: HgCl(2), TET and TlCl were more toxic in immature cultures, whereas BiNa-tartrate, dimethylmercury and NiCl(2) were more effective in differentiated cultures.