Patho- physiological role of BDNF in fibrin clotting

Circulating levels of Brain Derived Neurotrophic Factor (BDNF) are lower in coronary heart disease (CHD) than in healthy subjects and are associated with coronary events and mortality. However, the mechanism(s) underling this association is not fully understood. We hypothesize that BDNF may influence fibrin fiber structure and clot stability, favoring clot lysis and thrombus resolution. We showed that recombinant BDNF (rh-BDNF) influenced with clot formation in a concentration-dependent manner in both purified fibrinogen and plasma from healthy subjects. In particular, rh-BDNF reduced the density of fibrin fibers, the maximum clot firmness (MCF) and the maximum clot turbidity, and affected the lysis of clot. In addition, both thrombin and reptilase clotting time were prolonged by rh-BDNF, despite the amount of thrombin formed was greater. Intriguingly, CHD patients had lower levels of BDNF, greater fibrin fibers density, higher MCF than control subjects, and a negative correlation between BDNF and MCF was found. Of note, rh-BDNF markedly modified fibrin clot profile restoring physiological clot morphology in CHD plasma. In conclusion, we provide evidence that low levels of BDNF correlate with the formation of bigger thrombi (in vitro) and that this effect is mediated, at least partially, by the alteration of fibrin fibers formation

New insights into the biodegradation of thiodiglycol, the hydrolysis product of Yperite (sulfur mustard gas)

Aims: To isolate thiodiglycol (TDG)-degrading bacteria, the mustard gas hydrolysis product, and to characterize the metabolites formed and the enzymes involved in the degradation. Methods and Results: Two strains, identified as Achromobacter xylosoxydans G5 and Paracoccus denitrificans E4, isolated from a petroleum-contaminated soil, utilized TDG as sole carbon and sulfur source. During the degradation of TDG by strain E4 [(2-hydroxyethyl)thio] acetic acid (HETA), thiodiglycolic acid (TDGA) and bis-(2-hydroxyethyl)disulfide (BHEDS) were identified by gas chromatography\u2013mass spectrometry analysis, while HETA and TDGA were identified for strain G5. Two-dimensional isoelectric focussing-gel electrophoresis (2-D IEF/SDS\u2013PAGE) maps of protein extracts of P. denitrificans E4 grown on TDG showed a spot identified as a methanol dehydrogenase. Increased expression of a putative iscS gene, involved in sulfur assimilation, was observed in TDG-grown cells of A. xylosoxydans G5. Conclusions: TDG degradation by P. denitrificans E4 occurred through two pathways: one involved cleavage of the C\u2013S bond of HETA, yielding BHEDS and the other, oxidation of the alcoholic groups of TDG, yielding TDGA. The cleavage of the C\u2013S bond of TDGA gave mercaptoacetic acid, further oxidized to acetate and sulfate. Significance and Impact of the Study: Increased knowledge of TDG-degrading bacteria and the possibility of using them in a tailored-two-stage mustard gas destruction process

Identification of a putative LPS-associated cation exporter from Rhizobium leguminosarum bv. Viciae

A gene, cpaA, with similarity to calcium proton antiporters has been identified adjacent to lpcAB in Rhizobiumleguminosarum. LpcA is a galactosyl transferase while LpcB is a 2-keto-3-deoxyoctonate transferase, both of which are required to form the lipopolysaccharide (LPS) core in R. leguminosarum. Mutations in lpcAB result in a rough LPS phenotype with a requirement for elevated calcium concentrations to allow growth, suggesting that truncation of the LPS core exposes a highly negatively charged molecule. This is consistent with the LPS core being one of the main sites for binding calcium in the Gram-negative outer membrane. Strain RU1109 (cpaA::Tn5-lacZ) has a normal LPS layer, as measured by silver staining and Western blotting. This indicates that cpaA mutants are not grossly affected in their LPS layer. LacZ fusion analysis indicates that cpaA is constitutively expressed and is not directly regulated by the calcium concentration. Over-expression of cpaA increased the concentration of calcium required for growth, consistent with CpaA mediating calcium export from the cytosol. The location of lpcA, lpcB and cpaA as well as the phenotype of lpcB mutants suggests that CpaA might provide a specific export pathway for calcium to the LPS core

Assessing Free-Radical-Mediated DNA Damage during Cardiac Surgery : 8-Oxo-7,8-dihydro-2′-deoxyguanosine as a Putative Biomarker

Coronary artery bypass grafting (CABG), one of the most common cardiac surgical procedures, is characterized by a burst of oxidative stress. 8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), produced following DNA repairing, is used as an indicator of oxidative DNA damage in humans. The effect of CABG on oxidative-induced DNA damage, evaluated through the measurement of urinary 8-oxodG by a developed and validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method in 52 coronary artery disease (CAD) patients, was assessed before (T0), five days (T1), and six months (T2) after CABG procedure. These results were compared with those obtained in 40 subjects with cardiovascular risk factors and without overt cardiovascular disease (CTR). Baseline (T0) 8-oxodG was higher in CAD than in CTR (p = 0.035). A significant burst was detected at T1 (p = 0.019), while at T2, 8-oxodG levels were significantly lower than those measured at T0 (p < 0.0001) and comparable to those found in CTR (p = 0.73). A similar trend was observed for urinary 8-iso-prostaglandin F2\u3b1 (8-isoPGF2\u3b1 ), a reliable marker of oxidative stress. In the whole population baseline, 8-oxodG significantly correlated with 8-isoPGF2\u3b1 levels (r = 0.323, p = 0.002). These data argue for CABG procedure in CAD patients as inducing a short-term increase in oxidative DNA damage, as revealed by 8-oxodG concentrations, and a long-term return of such metabolite toward physiological levels

Nitric oxide synthetic pathway in red blood cells Is impaired in coronary artery disease

Background:All the enzymatic factors/cofactors involved in nitric oxide (NO) metabolism have been recently found in red blood cells. Increased oxidative stress impairs NO bioavailability and has been described in plasma of coronary artery disease (CAD) patients. The aim of the study was to highlight a potential dysfunction of the metabolic profile of NO in red blood cells and in plasma from CAD patients compared with healthy controls.Methods:We determined L-arginine/NO pathway by liquid-chromatography tandem mass spectrometry and high performance liquid chromatography methods. The ratio of oxidized and reduced forms of glutathione, as index of oxidative stress, was measured by liquid-chromatography tandem mass spectrometry method. NO synthase expression and activity were evaluated by immunofluorescence staining and ex-vivo experiments of L-[15N2]arginine conversion to L-[15N]citrulline respectively.Results:Increased amounts of asymmetric and symmetric dimethylarginines were found both in red blood cells and in plasma of CAD patients in respect to controls. Interestingly NO synthase expression and activity were reduced in CAD red blood cells. In contrast, oxidized/reduced glutathione ratio was increased in CAD and was associated to arginase activity.Conclusion:Our study analyzed for the first time the whole metabolic pathway of L-arginine/NO, both in red blood cells and in plasma, highlighting an impairment of NO pathway in erythrocytes from CAD patients, associated with decreased NO synthase expression/activity and increased oxidative stress

Gas Pixel Detectors for X-ray Polarimetry applications

We discuss a new class of Micro Pattern Gas Detectors, the Gas Pixel Detector (GPD), in which a complete integration between the gas amplification structure and the read-out electronics has been reached. An Application-Specific Integrated Circuit (ASIC) built in deep sub-micron technology has been developed to realize a monolithic device that is, at the same time, the pixelized charge collecting electrode and the amplifying, shaping and charge measuring front-end electronics. The CMOS chip has the top metal layer patterned in a matrix of 80 micron pitch hexagonal pixels, each of them directly connected to the underneath electronics chain which has been realized in the remaining five layers of the 0.35 micron VLSI technology. Results from tests of a first prototype of such detector with 2k pixels and a full scale version with 22k pixels are presented. The application of this device for Astronomical X-Ray Polarimetry is discussed. The experimental detector response to polarized and unpolarized X-ray radiation is shown. Results from a full MonteCarlo simulation for two astronomical sources, the Crab Nebula and the Hercules X1, are also reported.Comment: 16 pages, 20 figures, accepted for publication in Nuclear Instruments and Methods in Physics Research Section