Ascorbic acid reduces the frequency of iron induced micronuclei in bone marrow cells of mice

Iron is a potent oxidant that can lead to the formation of genotoxic lipid peroxides. Ascorbic acid, which enhances dietary iron absorption, has been suggested to enhance the oxidant effects of iron and to directly lead to the formation of lipid peroxides. The combined effects of dietary iron and ascorbic acid on genotoxicity were investigated by measuring the frequency of micronuclei in the bone marrow cells of C3H/He mice. In addition, liver iron concentration was measured in all treated groups. Three weeks old mice were fed diets for 3 weeks containing iron at 100 or 300 mg/kg diet in the form of FeSO₄ that were supplemented either with or without ascorbic acid (15 g/kg diet). The results of the bone marrow micronucleus test revealed that the high iron diet resulted in an increased frequency of micronucleated polychromatic erythrocytes (MnPCEs) as compared to low iron. Ascorbic acid supplementation in the low iron diet did not show any effect on incidence of MnPCEs and protected against the increased frequency of MnPCEs induced by the high iron diet. However, liver iron concentration was significantly increased only in the high iron treated and ascorbic acid supplemented group as compared to all other groups. These results demonstrate that ascorbic acid protects against the clastogenic effects of iron.     

ACUTE EFFECTS OF A RESISTED DYNAMIC WARM-UP PROTOCOL ON JUMPING PERFORMANCE

This study aimed to investigate the kinematic and kinetic changes when resistance is applied in horizontal and vertical directions, produced by using different percentages of body weight, caused by jumping movements during a dynamic warm-up. The group of subjects consisted of 35 voluntary male athletes (19 basketball and 16 volleyball players; age: 23.4 ± 1.4 years, training experience: 9.6 ± 2.7 years; height: 177.2 ± 5.7 cm, body weight: 69.9 ± 6.9 kg) studying Physical Education, who had a jump training background and who were training for 2 hours, on 4 days in a week. A dynamic warm-up protocol containing seven specific resistance movements with specific resistance corresponding to different percentages of body weight (2%, 4%, 6%, 8%, 10%) was applied randomly on non consecutive days. Effects of different warm-up protocols were assessed by pre-/post- exercise changes in jump height in the countermovement jump (CMJ) and the squat jump (SJ) measured using a force platform and changes in hip and knee joint angles at the end of the eccentric phase measured using a video camera. A significant increase in jump height was observed in the dynamic resistance warm-up conducted with different percentages of body weight (p < 0.05). On the other hand, no significant difference in different percentages of body weight states was observed (p > 0.05). In jump movements before and after the warm-up, while no significant difference between the vertical ground reaction forces applied by athletes was observed (p > 0.05), in some cases of resistance, a significant reduction was observed in hip and knee joint angles (p < 0.05). The dynamic resistance warm-up method was found to cause changes in the kinematics of jumping movements, as well as an increase in jump height values. As a result, dynamic warm-up exercises could be applicable in cases of resistance corresponding to 6-10% of body weight applied in horizontal and vertical directions in order to increase the jump performance acutely.     

Binpairs: Utilization of Illumina Paired-End Information for Improving Efficiency of Taxonomic Binning of Metagenomic Sequences

Motivation

Paired-end sequencing protocols, offered by next generation sequencing (NGS) platforms like Illumia, generate a pair of reads for every DNA fragment in a sample. Although this protocol has been utilized for several metagenomics studies, most taxonomic binning approaches classify each of the reads (forming a pair), independently. The present work explores some simple but effective strategies of utilizing pairing-information of Illumina short reads for improving the accuracy of taxonomic binning of metagenomic datasets. The strategies proposed can be used in conjunction with all genres of existing binning methods.

Results

Validation results suggest that employment of these “Binpairs” strategies can provide significant improvements in the binning outcome. The quality of the taxonomic assignments thus obtained are often comparable to those that can only be achieved with relatively longer reads obtained using other NGS platforms (such as Roche).

Availability

An implementation of the proposed strategies of utilizing pairing information is freely available for academic users at https://metagenomics.atc.tcs.com/binning/binpairs.     

Activation characteristics of transient receptor potential ankyrin 1 and its role in nociception

Transient receptor potential (TRP) ankyrin 1 (TRPA1) is a Ca(2+)-permeant, nonselective cationic channel. It is predominantly expressed in the C afferent sensory nerve fibers of trigeminal and dorsal root ganglion neurons and is highly coexpressed with the nociceptive ion channel transient receptor potential vanilloid 1 (TRPV1). Several physical and chemical stimuli have been shown to activate the channel. In this study, we have used electrophysiological techniques and behavioral models to characterize the properties of TRPA1. Whole cell TRPA1 currents induced by brief application of lower concentrations of N-methyl maleimide (NMM) or allyl isothiocyanate (AITC) can be reversed readily by washout, whereas continuous application of higher concentrations of NMM or AITC completely desensitized the currents. The deactivation and desensitization kinetics differed between NMM and AITC. TRPA1 current amplitude increased with repeated application of lower concentrations of AITC, whereas saturating concentrations of AITC induced tachyphylaxis, which was more pronounced in the presence of extracellular Ca(2+). The outward rectification exhibited by native TRPA1-mediated whole cell and single-channel currents was minimal as compared with other TRP channels. TRPA1 currents were negatively modulated by protons and polyamines, both of which activate the heat-sensitive channel, TRPV1. Interestingly, neither protein kinase C nor protein kinase A activation sensitized AITC-induced currents, but each profoundly sensitized capsaicin-induced currents. Current-clamp experiments revealed that AITC produced a slow and sustained depolarization as compared with capsaicin. TRPA1 is also expressed at the central terminals of nociceptors at the caudal spinal trigeminal nucleus. Activation of TRPA1 in this area increases the frequency and amplitude of miniature excitatory or inhibitory postsynaptic currents. In behavioral studies, intraplantar and intrathecal administration of AITC induced more pronounced and prolonged changes in nociceptive behavior than those induced by capsaicin. In conclusion, the characteristics of TRPA1 we have delineated suggest that it might play a unique role in nociception.     

Four structural subclasses of the antivirulence drug target disulfide oxidoreductase DsbA provide a platform for design of subclass-specific inhibitors

By catalyzing oxidative protein folding, the bacterial disulfide bond protein A (DsbA) plays an essential role in the assembly of many virulence factors. Predictably, DsbA disruption affects multiple downstream effector molecules, resulting in pleiotropic effects on the virulence of important human pathogens. These findings mark DsbA as a master regulator of virulence, and identify the enzyme as a target for a new class of antivirulence agents that disarm pathogenic bacteria rather than killing them. The purpose of this article is to discuss and expand upon recent findings on DsbA and to provide additional novel insights into the druggability of this important disulfide oxidoreductase by comparing the structures and properties of 13 well-characterized DsbA enzymes. Our structural analysis involved comparison of the overall fold, the surface properties, the conformations of three loops contributing to the binding surface and the sequence identity of residues contributing to these loops. Two distinct structural classes were identified, classes I and II, which are differentiated by their central β-sheet arrangements and which roughly separate the DsbAs produced by Gram-negative from Gram-positive organisms. The classes can be further subdivided into a total of four subclasses on the basis of surface features. Class Ia is equivalent to the Enterobacteriaceae class that has been defined previously. Bioinformatic analyses support the classification of DsbAs into 3 of the 4 subclasses, but did not pick up the 4th subclass which is only apparent from analysis of DsbA electrostatic surface properties. In the context of inhibitor development, the discrete structural subclasses provide a platform for developing DsbA inhibitory scaffolds with a subclass-wide spectrum of activity. We expect that more DsbA classes are likely to be identified, as enzymes from other pathogens are explored, and we highlight the issues associated with structure-based inhibitor development targeting this pivotal mediator of bacterial virulence. This article is part of a Special Issue entitled: Thiol-Based Redox Processes.     

Mutation at I-A beta chain prevents experimental autoimmune myasthenia gravis

Immune response (Ir) gene(s) at the I-A subregion of the mouse H-2 complex influence susceptibility to experimental autoimmune myasthenia gravis (EAMG). To determine the importance of the Ir gene product, the Ia antigens, in EAMG pathogenesis, we studied the degree of EAMG susceptibility of an I-A mutant strain, the B6.C-H-2 ^bm12 (bm12), and its parent B6/Kh. According to the cellular, humoral, biochemical, and clinical manifestations of EAMG, the I-A mutation converted an EAMG susceptible strain (B6/Kh) into a relatively resistant strain (bm12). The relative resistance to EAMG induction in bm12 may be due to the lack of Ia.8 and/or la.39 determinants and/or quantitative expression of la antigens.Abbreviations used in this paper MG myasthenia gravis - AChR acetylcholine receptors - EAMG experimental autoimmune myasthenia gravis - Ir immune response - B6 C57BL/6J - bm12 B6.C-H-2 ^bm12 - CFA complete Freund's adjuvant - LNC lymph node cells - PPD purified protein derivative     

Muscarinic acetylcholine receptor produced in recombinant baculovirus infected Sf9 insect cells couples with endogenous G-proteins to activate ion channels.

Following the infection of insect ovarian cells (Sf9) with recombinant bearing the cDNA coding for the rat muscarinic acetylcholine (ACh) receptor subtype m3, ionic flux across the membrane in response to the application of ACh was examined electrophysiologically. We show that ACh activates potassium currents. The response is abolished when cells are treated with pertussis toxin. No ACh-induced currents are observed from uninfected cells or cells infected with virus which do not contain the cDNA coding for ACh receptors in its genome. The characteristics of single channel currents show time-dependent changes following the application of ACh. Initially, ACh activates brief channel currents with a conductance of about 5 pS. The conductance level of channels gradually increases in steps to 10 pS and then to 20 pS and 40 pS. At the same time, channel open probability also increases. Thereafter, additional channels appear, opening and closing independently of, or at times in synchrony with, the original channel.     

Stoichiometry of Recombinant N-Methyl-d-Aspartate Receptor Channels Inferred from Single-channel Current Patterns

Single-channel currents were recorded from mouse NR1-NR2B (ζ-ε₂) receptors containing mixtures of wild-type and mutant subunits expressed in Xenopus oocytes. Mutant subunits had an asparagine-to-glutamine (N-to-Q) mutation at the N₀ site of the M2 segment (NR1:598, NR2B:589). Receptors with pure N or Q NR1 and NR2 subunits generated single-channel currents with distinctive current patterns. Based on main and sublevel amplitudes, occupancy probabilities, and lifetimes, four patterns of current were identified, corresponding to receptors with the following subunit compositions (NR1/NR2): N/N, N/Q, Q/N, and Q/Q. Only one current pattern was apparent for each composition. When a mixture of N and Q NR2 subunits was coexpressed with pure mutant NR1 subunits, three single-channel current patterns were apparent. One pattern was the same as Q/Q receptors and another was the same as Q/N receptors. The third, novel pattern presumably arose from hybrid receptors having both N and Q NR2 subunits. When a mixture of N and Q NR1 subunits was coexpressed with pure mutant NR2 subunits, six single-channel current patterns were apparent. One pattern was the same as Q/Q receptors and another was the same as N/Q receptors. The four novel patterns presumably arose from hybrid receptors having both N and Q NR1 subunits. The relative frequency of NR1 hybrid receptor current patterns depended on the relative amounts of Q and N subunits that were injected into the oocytes. The number of hybrid receptor patterns suggests that there are two NR2 subunits per receptor and is consistent with either three or five NR1 subunits per receptor, depending on whether or not the order of mutant and wild-type subunits influences the current pattern. When considered in relation to other studies, the most straightforward interpretation of the results is that N-methyl-d-aspartate receptors are pentamers composed of three NR1 and two NR2 subunits.