Methylenedioxymethamphetamine (Ecstasy) Decreases Neutrophil Activity and Alters Leukocyte Distribution in Bone Marrow, Spleen and Blood

Objective: Looking for possible neuroimmune relationships, we analyzed the effects of methylenedioxymethamphetamine (MDMA) administration on neuroendocrine, neutrophil activity and leukocyte distribution in mice. Methods: Five experiments were performed. In the first, mice were treated with MDMA (10 mg/kg) 30, 60 min and 24 h prior to blood sample collection for neutrophil activity analysis. In the second experiment, the blood of nave mice was collected and incubated with MDMA for neutrophil activity in vitro analysis. In the third and fourth experiments, mice were injected with MDMA (10 mg/kg) and 60 min later, blood and brain were collected to analyze corticosterone serum levels and hypothalamic noradrenaline (NA) levels and turnover. In the last experiment, mice were injected with MDMA 10 mg/kg and 60 min later, blood, bone marrow and spleen were collected for leukocyte distribution analysis. Results: Results showed an increase in hypothalamic NA turnover and corticosterone serum levels 60 min after MDMA (10 mg/kg) administration, a decrease in peripheral blood neutrophil oxidative burst and a decrease in the percentage and intensity of neutrophil phagocytosis. It was further found that MDMA (10 mg/kg) treatment also altered leukocyte distribution in blood, bone marrow and spleen. In addition, no effects were observed for MDMA after in vitro exposure both in neutrophil oxidative burst and phagocytosis. Conclusion: The effects of MDMA administration (10 mg/kg) on neutrophil activity and leukocyte distribution might have been induced indirectly through noradrenergic neurons and/or hypothalamic-pituitary-adrenal axis activations. Copyright (C) 2009 S. Karger AG, Basel

Transcriptional responses of host peripheral blood cells to tuberculosis infection

Host responses following exposure to Mycobacterium tuberculosis (TB) are complex and can significantly affect clinical outcome. These responses, which are largely mediated by complex immune mechanisms involving peripheral blood cells (PBCs) such as T-lymphocytes, NK cells and monocyte-derived macrophages, have not been fully characterized. We hypothesize that different clinical outcome following TB exposure will be uniquely reflected in host gene expression profiles, and expression profiling of PBCs can be used to discriminate between different TB infectious outcomes. In this study, microarray analysis was performed on PBCs from three TB groups (BCG-vaccinated, latent TB infection, and active TB infection) and a control healthy group. Supervised learning algorithms were used to identify signature genomic responses that differentiate among group samples. Gene Set Enrichment Analysis was used to determine sets of genes that were co-regulated. Multivariate permutation analysis (p < 0.01) gave 645 genes differentially expressed among the four groups, with both distinct and common patterns of gene expression observed for each group. A 127-probeset, representing 77 known genes, capable of accurately classifying samples into their respective groups was identified. In addition, 13 insulin-sensitive genes were found to be differentially regulated in all three TB infected groups, underscoring the functional association between insulin signaling pathway and TB infection. Published by Elsevier Ltd.

Biomonitoring method for the simultaneous determination of cadmium and lead in whole blood by electrothermal atomic absorption spectrometry for assessment of environmental exposure

The aim of this work is to propose a biomonitoring method for the simultaneous determination of Cd and Pb in whole blood by simultaneous electrothermal atomic absorption spectrometry for assessment of environmental levels. A volume of 200 mu L of whole blood was diluted in 500 mu L of 0.2% (w v(-1)) Triton(R) X-100 + 2.0% (v v(-1)) HNO3. Trichloroacetic acid was added for protein precipitation and the supernatant analyzed. A mixture of 250 mu g W + 200 mu g Rh as permanent and 2.0% (w v(-1)) NH4H2PO4 as co-injected modifiers were used. Characteristic masses and limits of detections (n = 20, 3s) for Cd and Pb were 1.26 and 33 pg and 0.026 mu g L-1 and 0.65 mu g L-1, respectively. Repeatability ranged from 1.8 to 6.8% for Cd and 1.2 to 1.7% for Pb. The trueness of method was checked by the analysis of three Reference Materials: Lyphocheck(R) Whole Blood Metals Control level 1 and Seronorm(TM) Trace Elements in Whole Blood levels 1 and 2. The found concentrations presented no statistical differences at the 95% confidence level. Blood samples from 40 volunteers without occupational exposure were analyzed and the concentrations ranged from 0.13 to 0.71 mu g L-1 (0.32 +/- 0.19 mu g L-1) for Cd and 9.3 to 56.7 mu g L-1 (25.1 +/- 10.8 mu g L-1) for Pb. (C) 2007 Elsevier B.V. All rights reserved.

Blood pressure variability increases connexin expression in the vascular smooth muscle of rats

Aims Following sinoaortic denervation (SAD), isolated rat aortas present oscillatory contractions and demonstrate a heightened contraction for alpha-adrenergic agonists. Our aim was to verify the effects of SAD on connexin43 (Cx43) expression and phenylephrine-induced contraction in isolated aortas. Methods and results Three days after surgery (SAD or sham operation), isolated aortic rings were exposed to phenylephrine and acetylcholine (0.1-10 mu M) in the presence or absence of the gap junction blocker 18 beta-glycyrrhetinic acid (18 beta-GA, 100 mu M). Vascular reactivity to potassium chloride (KCl, 4.7-120 mM) was also examined. The incidence of rats presenting oscillatory contractions was measured. Effects of SAD on the vascular smooth muscle expression of the Cx43 mRNA by RT-PCR and western blotting for Cx43 protein were examined. Phenylephrine-induced contraction was higher in SAD rat aortas compared with the control. In the presence of 18 beta-GA, the response to phenylephrine was similar in both groups. Oscillatory contractions were observed in 10/10 SAD rat aortas vs. 2/10 controls. Relaxing response to acetylcholine was similar in both groups, but in the presence of 18 beta-GA, the response to acetylcholine decreased significantly in the sham-operated group (82.7 +/- 7.6% reduction of relaxation), whereas a half-maximal relaxation (reduction of 46.2 +/- 5.3%) took place in SAD rat aortas. KCl-induced contraction was similar in both groups. Following SAD, RT-PCR revealed significantly increased levels of Cx43 mRNA (9.85 fold, P < 0.01). Western blot analysis revealed greater levels of Cx43 protein (P < 0.05). Conclusion Blood pressure variability evoked by SAD leads to increased expression of Cx43, which could contribute to enhanced phenylephrine-induced contraction and oscillatory activity in isolated aortas.

Liquid-phase microextraction combined with high-performance liquid chromatography for the enantioselective analysis of mefloquine in plasma samples

A simple and rapid method, which involves liquid-phase microextraction (LPME) followed by HPLC analysis using Chiralpak AD column and UV detection, was developed for the enantioselective determination of mefloquine in plasma samples. Several factors that influence the efficiency of three-phase LPME were investigated and optimized. Under the optimal extraction conditions, the mean recoveries were 33.2 and 35.0% for (-)-(SR-)-mefloquine and (+)-(RS)-mefloquine, respectively. The method was linear over 50-1500 ng/ml range. Within-day and between-day assay precision and accuracy were below 15% for both enantiomers at concentrations of 150, 600 and 1200 ng/ml. Furthermore, no racemization or degradation were seen with the method described. (C) 2007 Elsevier B.V. All rights reserved.

Methylmercury and inorganic mercury determination in blood by using liquid chromatography with inductively coupled plasma mass spectrometry and a fast sample preparation procedure

Despite the necessity to differentiate chemical species of mercury in clinical specimens, there area limited number of methods for this purpose. Then, this paper describes a simple method for the determination of methylmercury and inorganic mercury in blood by using liquid chromatography with inductively coupled mass spectrometry (LC-ICP-MS) and a fast sample preparation procedure. Prior to analysis, blood (250 mu L) is accurately weighed into 15-mL conical tubes. Then, an extractant solution containing mercaptoethanol, L-cysteine and HCI was added to the samples following sonication for 15 min. Quantitative mercury extraction was achieved with the proposed procedure. Separation of mercury species was accomplished in less than 5 min on a C18 reverse-phase column with a mobile phase containing 0.05% (v/v) mercaptoethanol, 0.4% (m/v) L-cysteine, 0.06 mol L(-1) ammonium acetate and 5% (v/v) methanol. The method detection limits were found to be 0.25 mu g L(-1) and 0.1 mu Lg L(-1) for inorganic mercury and methylmercury, respectively. Method accuracy is traceable to Standard Reference Material (SRM) 966 Toxic Metals in Bovine Blood from the National Institute of Standards and Technology (NIST). The proposed method was also applied to the speciation of mercury in blood samples collected from fish-eating communities and from rats exposed to thimerosal. With the proposed method there is a considerable reduction of the time of sample preparation prior to speciation of Hg by LC-ICP-MS. Finally, after the application of the proposed method, we demonstrated an interesting in vivo ethylmercury conversion to inorganic mercury. (C) 2009 Elsevier B.V. All rights reserved.

Elevated blood lead levels in a riverside population in the Brazilian Amazon

Lead (Pb) is recognized as one of the most toxic metals. Sources of Pb exposure have been widely documented in North America, and the removal of Pb additives from gasoline was reflected in a dramatic lowering of blood Pb concentration. In Latin America, the removal of Pb from gasoline resulted in decreased exposure, but Pb levels in many areas remain high due to occupational and environmental sources of exposure. While many of the Pb sources have been identified (mining, industries, battery recycling, lead-based paint, ceramics), new ones occasionally crop up. Here we report on blood Pb (B-Pb) levels in remote riverside communities of the Brazilian Amazon. Blood Pb (B-Pb) levels were determined in 448 persons from 12 villages of the Lower Tapajos River Basin, Par, Brazil. Socio-demographic and dietary information, as well as occupational, residential and medical history was collected using an interview-administered questionnaire. B-Pb, measured by ICP-MS, showed elevated concentrations. Mean B-Pb was 13.1 mu g/dL +/- 8.5, median B-Pb was 11.2 mu g/dL and ranged from 0.59 to 48.3 mu g/dL. Men had higher B-Pb compared to women (median: 15.3 mu g/dL vs 7.9 mu g/dL respectively). B-Pb increased with age for women, while it decreased for men. For both genders, B-Pb decreased with education. There were significant differences between villages. Exploratory analyses, using linear partition models, showed that for men B-Pb was lower among those who were involved in cattle-raising, and higher among those who hunted, farmed and fished. The distribution profile of B-Pb directed us towards artisanal transformation of manioc to flour (farinha), which requires heating in a large metal pan, with stirring primarily done by young men. In the village with the highest B-Pb, analysis of Pb concentrations (dry weight) of manioc (prior to transformation) and farinha (following transformation) from 6 houses showed a tenfold increase in Pb concentration (mean: 0.017 +/- 0.016 to 0.19 +/- 0.10 mu g/g). This was confirmed in one of these villages where we sampled manioc paste Oust before roasting) and the roasted farinha (0.05 mu g/g vs 0.20 mu g/g). While there may be other sources (ammunition, sinkers for fishing nets), the high concentrations in farinha, a dietary staple, assuredly makes an important contribution. Further action needs to reduce Pb sources in this region. (C) 2009 Elsevier Inc. All rights reserved.

Mercury speciation in whole blood by gas chromatography coupled to ICP-MS with a fast microwave-assisted sample preparation procedure

A simple and fast method is described for simultaneous determination of methylmercury (MeHg), ethylmercury (Et-Hg) and inorganic mercury (Ino-Hg) in blood samples by using capillary gas chromatography-inductively coupled plasma mass spectrometry (GC-ICP-MS) after derivatization and alkaline digestion. Closed-vessel microwave assisted digestion conditions with tetramethylammonium hydroxide (TMAH) have been optimized. Derivatization by using ethylation and propylation procedures have also been evaluated and compared. The absolute detection limits (using a 1 mu L injection) obtained by GC-ICP-MS with ethylation were 40 fg for MeHg and Ino-Hg, respectively, and with propylation were 50, 20 and 50 fg for MeHg, Et-Hg and Ino-Hg, respectively. Method accuracy is traceable to Standard Reference Material (SRM) 966 Toxic Metals in Bovine Blood from the National Institute of Standards and Technology (NIST). Additional validation is provided based on the comparison of results obtained for mercury speciation in blood samples with the proposed procedure and with a previously reported LC-ICP-MS method. With the new proposed procedure no tedious clean-up steps are required and a considerable improvement of the time of analysis was achieved compared to other methods using GC separation.

Stereoselective analysis of carvedilol in human plasma and urine using HPLC after chiral derivatization

An enantioselective high-performance liquid chromatographic method for the analysis of carvedilol in plasma and urine was developed and validated using (-)-menthyl chloroformate (MCF) as a derivatizing reagent. Chloroform was used for extraction, and analysis was performed by HPLC on a C18 column with a fluorescence detector. The quantitation limit was 0.25 ng/ml for S(-)-carvedilol in plasma and 0.5 ng/ml for R(+)-carvedilol in plasma and for both enantiomers in urine. The method was applied to the study of enantioselectivity in the pharmacokinetics of carvedilol administered in a multiple dose regimen (25mg/12h) to a hypertensive elderly female patient. The data obtained demonstrated highest plasma levels for the R(+)-carvedilol(AUCSS 75.64 vs 37.29ng/ml). The enantiomeric ratio R(+)/S(-) was 2.03 for plasma and 1.49 0 - 12 for urine (Aeo-12 17.4 vs 11.7 pg). Copyright (c) 2008 John Wiley & Sons, Ltd.

Simultaneous Analysis of Tramadol, O-Desmethyltramadol, and N-Desmethyltramadol Enantiomers in Rat Plasma by High-Performance Liquid Chromatography-Tandem Mass Spectrometry: Application to Pharmacokinetics

Tramadol (T) is available as a racemic mixture of (+)-trans-T and (-)-trans-T. The main metabolic pathways are O-demethylation and N-demethylation, producing trans-O-desmethyltramadol (M1) and trans-N-desmethyltramadol (M2) enantiomers, respectively. The analgesic effect of T is related to the opioid activity of (+)-trans-T and (+)-M1 and to the monoaminergic action of (+/-)-trans-T. This is the first study using tandem mass spectrometry as a detection system for the simultaneous analysis of trans-T, M1, and M2 enantiomers. The analytes were resolved on a Chiralpak (R) AD column using hexane: ethanol (95.5:4.5, v/v) plus 0.1% diethylamine as the mobile phase. The quantitation limits were 0.5 ng/ml for trans-T and M1 and 0.1 ng/ml for M2. The method developed and validated here was applied to a pharmacokinetic study in rats. Male Wistar rats (n = 6 at each time point) received a single oral dose of 20 mg/kg racemic trans-T. Blood samples were collected up to 12 h after drug administration. The kinetic disposition of trans-T and M2 was enantioselective (AUC((+)/(-)) ratio = 4.16 and 6.36, respectively). The direction and extent of enantioselectivity in the pharmacokinetics of trans-T and M2 in rats were comparable to data previously reported for healthy volunteers, suggesting that rats are a suitable model for enantioselective studies of trans-T pharmacokinetics. Chirality 23: 287-293, 2011. (C) 2010 Wiley-Liss, Inc.

Biomarkers of selenium status in the amazonian context: Blood, urine and sequential hair segments

Selenium (Se) is an essential element and deficit or excess of dietary Se is associated with health disorders. Relatively elevated Se levels have been reported in the Brazilian Amazon, where there are also important annual variations in the availability of different foods. The present study was conducted among six riparian communities of the Tapajos River to evaluate seasonal variations in blood and sequential hair cm Se concentrations, and to examine the relationships between Se in blood and hair, and blood and urine. Two cross-sectional studies were conducted, at the descending water (DWS, n = 259) and the rising water (RWS, n = 137) seasons, with repeated measures for a subgroup (n = 112). Blood Se (B-Se), hair Se (H-Se) and urine Se (U-Se) were determined. Match-paired analyses were used for seasonal comparisons and the method of best fit was used to describe the relationships between biomarkers. B-Se levels presented a very large range (142-2447 mu g/l) with no overall seasonal variation (median 284 and 292 mu g/l, respectively). Sequential analysis of 13 cm hair strands showed significant variations over time: Se concentrations at the DWS were significantly lower compared with the rising water season (medians: 0.7 and 0.9 mu g/g; ranges: 0.2-4.3 mu g/g and 0.2-5.4 mu g/g, respectively). At both seasons, the relationships between B-Se and H-Se were linear and highly significant (r(2) = 67.9 and 63.6, respectively), while the relationship between B-Se and U-Se was best described by a sigmoid curve. Gender, age, education and smoking did not influence Se status or biomarker relationships. Variations in H-Se suggest that there may be seasonal availability of Se sources in local food. For populations presenting a large range and/or elevated Se exposure, sequential analyses of H-Se may provide a good reflection of variations in Se status.

Analysis of tissue responses to fin tagging in Australian carcharhinids

Tissue responses to the application of Rototags and Jumbo Rototags in the first dorsal fin of Carcharhinus melanopterus, C. obscurus and C. plumbeus were examined. The acute response included tissue tearing and haemorrhage and was present by 5 days post-tagging. The intermediate response had begun by 20 days post-tagging and continued beyond 207 days. This response involved decreased red blood cell activity as the inflammatory response commenced. The chronic response had begun by 301 days and was complete by 553 days with a layer of fibrous connective tissue walling off the tag. External damage to the fin was caused by continued abrasion by the tag. Repair scales were observed at 242 days using scanning electron microscopy and were confirmed histologically in 61- and 553-day samples. Repair scales were not seen in areas of continuous abrasion. No infection was observed in tissues surrounding the wound. Disruption of the fin surface was observed due to abrasion by the tag, but did not appear to cause a severe tissue reaction. The tissue responses observed were consistent with a normal, but relatively slow, healing in the vicinity of the tag wound. Use of Rototags or Jumbo Rototags appears to be an efficient way of marking elasmobranchs with minimal damage to the shark. (C) 1998 The Fisheries Society of the British Isles.

Heart rate, blood lactate and kinematic data of elite colts (under-19) rugby union players during competition

Physiological and kinematic data were collected from elite under-19 rugby union players to provide a greater understanding of the physical demands of rugby union. Heart rate, blood lactate and time-motion analysis data were collected from 24 players (mean +/- s((x) over bar): body mass 88.7 +/- 9.9 kg, height 185 +/- 7 cm, age 18.4 +/- 0.5 years) during six competitive premiership fixtures. Six players were chosen at random from each of four groups: props and locks, back row forwards, inside backs, outside backs. Heart rate records were classified based on percent time spent in four zones (>95%, 85-95%, 75-84%, <75% HRmax). Blood lactate concentration was measured periodically throughout each match, with movements being classified as standing, walking, jogging, cruising, sprinting, utility, rucking/mauling and scrummaging. The heart rate data indicated that props and locks (58.4%) and back row forwards (56.2%) spent significantly more time in high exertion (85-95% HRmax) than inside backs (40.5%) and outside backs (33.9%) (P < 0.001). Inside backs (36.5%) and outside backs (38.5%) spent significantly more time in moderate exertion (75-84% HRmax) than props and locks (22.6%) and back row forwards (19.8%) (P < 0.05). Outside backs (20.1%) spent significantly more time in low exertion (< 75% HRmax) than props and locks (5.8%) and back row forwards (5.6%) (P < 0.05). Mean blood lactate concentration did not differ significantly between groups (range: 4.67 mmol.l(-1) for outside backs to 7.22 mmol.l(-1) for back row forwards; P < 0.05). The motion analysis data indicated that outside backs (5750 m) covered a significantly greater total distance than either props and locks or back row forwards (4400 and 4080 m, respectively; P < 0.05). Inside backs and outside backs covered significantly greater distances walking (1740 and 1780 m, respectively; P < 0.001), in utility movements (417 and 475 m, respectively; P < 0.001) and sprinting (208 and 340 m, respectively; P < 0.001) than either props and locks or back row forwards (walking: 1000 and 991 m; utility movements: 106 and 154 m; sprinting: 72 and 94 m, respectively). Outside backs covered a significantly greater distance sprinting than inside backs (208 and 340 m, respectively; P < 0.001). Forwards maintained a higher level of exertion than backs, due to more constant motion and a large involvement in static high-intensity activities. A mean blood lactate concentration of 4.8-7.2 mmol.l(-1) indicated a need for 'lactate tolerance' training to improve hydrogen ion buffering and facilitate removal following high-intensity efforts. Furthermore, the large distances (4.2-5.6 km) covered during, and intermittent nature of, match-play indicated a need for sound aerobic conditioning in all groups (particularly backs) to minimize fatigue and facilitate recovery between high-intensity efforts.

Bioelectrical impedance analysis predicts outcome in patients with suspected bacteremia

Fluid shifts from intracellular to extracellular water (ICW to ECW) are a feature of sepsis, caused by increased vascular permeability and cell catabolism. Changes in ECW and total body water (TBW) were assessed in a prospective observational study of patients with bacteremia by a bedside technique, and its prognostic impact determined; In 78 hospital patients with fever, the resistance ratio (Rinf/RO) and estimated ECW/TBW ratio from multifrequency bioelectrical impedance analysis, and serum albumin concentration were measured. Rinf/RO and ECW/TBW ratios decreased from day 0 to 2 in patients with significant bacteremia (n = 31), but not in patients with doubtful or negative blood cultures (n = 22 and 25), Increased Rinf/RO at baseline, and further increase of ECW/TBW from day 0 to 2, were associated with lower rate of recovery after 1 week and with higher mortality. Baseline Rinf/RO above the median (0.75) had positive and negative predictive values of 0.31 and 0.95 for death. This prognostic effect was independent of underlying disease and blood culture result in a multivariate model. Hypoalbuminemia at baseline was predictive of outcome, but changes in albumin from day 0 to 2 were unrelated to blood culture results or outcome. In patients with bacteremia,fluid shifts from intracellular to extracellular,vater occur early are rapidly reversible by antibiotic treatment but are associated with adverse prognosis. Bioelectrical impedance deserves further study as a tool for bedside monitoring of patients with bacteremia.

Quantitative analysis of vascular to cardiac selectivity of L- and T-type voltage-operated calcium channel antagonists in human tissues

1. Classical L-type voltage-operated calcium channel (VOCC) antagonists dilate blood vessels, depress myocardial contractility and slow cardiac conduction. 2. We compared four L-type VOCC antagonists and a novel tetralol derivative, mibefradil, reportedly 10-fold more selective for T- (transient) over L-type VOCC in two in vitro assays of human tissue, namely isolated small arteries from the aortic vasa vasorum in a myograph and right atrial trabeculae muscle under isometric force conditions. 3. In arteries contracted with K+ (62 mmol/L), the relaxation pIC(50) values for the VOCC antagonists felodipine, nifedipine, amlodipine, verapamil and mibefradil were 8.30, 7.78, 6.64, 6.26 and 6.22, respectively. In atrial trabeculae, the pIC(50) values to inhibit the inotropic response to a submaximal concentration of isoprenaline (6 nmol/L) for felodipine, nifedipine, verapamil, amlodipine and mibefradil were 7.21, 6.95, 6.91, 5.94 and 4.61, respectively. 4. Taking the anti-log (pIC(50) vessel - pIC(50) atrium) the vascular relaxation to cardiac depression potency ratios for mibefradil, felodipine, nifedipine, amlodipine and verapamil were 41, 12, 7, 5 and 0.22, respectively. 5. We conclude that, in human tissue assays, perhaps T- over L-type VOCC selectivity confers the most favourable vascular selectivity on mibefradil. Alternatively, splice variants of L-type VOCC in the vasculature (CaV1.2b) may be more sensitive to mibefradil than the splice variants in the heart (CaV1.2a).