Genotoxicity investigation of ELF-magnetic fields in Salmonella typhimurium with the sensitive SOS-based VITOTOX test

We performed a genotoxicity investigation of extremely low-frequency (ELF) magnetic fields (MFs, 50 Hz, 100 and 500 μT, 1 and 2 h exposure) alone and in combination with known chemical mutagens using the VITOTOX test. This test is a very sensitive reporter assay of Salmonella typhimurium bacteria based on the SOS response. Our study showed that ELF-MFs do not induce SOS-based mutagenicity in S. typhimurium bacteria and do not show any synergetic effect when combined with chemical mutagens.     

Mathematical modelling of the mitochondrial apoptosis pathway

Mitochondria are pivotal for cellular bioenergetics, but are also a core component of the cell death machinery. Hypothesis-driven research approaches have greatly advanced our understanding of the role of mitochondria in cell death and cell survival, but traditionally focus on a single gene or specific signalling pathway at a time. Predictions originating from these approaches become limited when signalling pathways show increased complexity and invariably include redundancies, feedback loops, anisotropies or compartmentalisation. By introducing methods from theoretical chemistry, control theory, and biophysics, computational models have provided new quantitative insights into cell decision processes and have led to an increased understanding of the key regulatory principles of apoptosis. In this review, we describe the currently applied modelling approaches, discuss the suitability of different modelling techniques, and evaluate their contribution to the understanding of the mitochondrial apoptosis pathway. This article is part of a Special Issue entitled Mitochondria: the deadly organelle.     

Cell culture replication of a genotype 1b hepatitis C virus isolate cloned from a patient who underwent liver transplantation

The introduction of the genotype 2a isolate JFH1 was a major breakthrough in the field of hepatitis C virus (HCV), allowing researchers to study the complete life cycle of the virus in cell culture. However, fully competent culture systems encompassing the most therapeutically relevant HCV genotypes are still lacking, especially for the highly drug-resistant genotype 1b. For most isolated HCV clones, efficient replication in cultured hepatoma cells requires the introduction of replication-enhancing mutations. However, such mutations may interfere with viral assembly, as occurs in the case of the genotype 1b isolate Con1. In this study, we show that a clinical serum carrying a genotype 1b virus with an exceptionally high viral load was able to infect Huh7.5 cells. Similar to previous reports, inoculation of Huh7.5 cells by natural virus is very inefficient compared to infection by cell culture HCV. A consensus sequence of a new genotype 1b HCV isolate was cloned from the clinical serum (designated Barcelona HCV1), and then subjected to replication studies. This virus replicated poorly in a transient fashion in Huh7.5 cells after electroporation with in vitro transcribed RNA. Nonetheless, approximately 3 weeks post electroporation and thereafter, core protein-positive cells were detected by immunofluorescence. Surprisingly, small amounts of core protein were also measurable in the supernatant of electroporated cells, suggesting that HCV particles might be assembled and released. Our findings not only enhance the current method of cloning in vitro HCV replication-competent isolates, but also offer valuable insights for the realization of fully competent culture systems for HCV.     

Subtype differences in pre-coupling of muscarinic acetylcholine receptors

Based on the kinetics of interaction between a receptor and G-protein, a myriad of possibilities may result. Two extreme cases are represented by: 1/Collision coupling, where an agonist binds to the free receptor and then the agonist-receptor complex "collides" with the free G-protein. 2/Pre-coupling, where stable receptor/G-protein complexes exist in the absence of agonist. Pre-coupling plays an important role in the kinetics of signal transduction. Odd-numbered muscarinic acetylcholine receptors preferentially couple to G(q/11), while even-numbered receptors prefer coupling to G(i/o). We analyzed the coupling status of the various subtypes of muscarinic receptors with preferential and non-preferential G-proteins. The magnitude of receptor-G-protein coupling was determined by the proportion of receptors existing in the agonist high-affinity binding conformation. Antibodies directed against the C-terminus of the α-subunits of the individual G-proteins were used to interfere with receptor-G-protein coupling. Effects of mutations and expression level on receptor-G-protein coupling were also investigated. Tested agonists displayed biphasic competition curves with the antagonist [(3)H]-N-methylscopolamine. Antibodies directed against the C-terminus of the α-subunits of the preferential G-protein decreased the proportion of high-affinity sites, and mutations at the receptor-G-protein interface abolished agonist high-affinity binding. In contrast, mutations that prevent receptor activation had no effect. Expression level of preferential G-proteins had no effect on pre-coupling to non-preferential G-proteins. Our data show that all subtypes of muscarinic receptors pre-couple with their preferential classes of G-proteins, but only M(1) and M(3) receptors also pre-couple with non-preferential G(i/o) G-proteins. Pre-coupling is not dependent on agonist efficacy nor on receptor activation. The ultimate mode of coupling is therefore dictated by a combination of the receptor subtype and the class of G-protein.     

Lysate of probiotic Lactobacillus casei DN-114 001 ameliorates colitis by strengthening the gut barrier function and changing the gut microenvironment

Background

Probiotic bacteria can be used for the prevention and treatment of human inflammatory diseases including inflammatory bowel diseases (IBD). However, the nature of active components and exact mechanisms of this beneficial effects have not been fully elucidated. Our aim was to investigate if lysate of probiotic bacterium L. casei DN-114 001 (Lc) could decrease the severity of intestinal inflammation in a murine model of IBD.

Methodology/Principal Findings

The preventive effect of oral administration of Lc significantly reduces the severity of acute dextran sulfate sodium (DSS) colitis in BALB/c but not in SCID mice. In order to analyze how this beneficial effect interferes with well-known phases of intestinal inflammation pathogenesis in vivo and in vitro, we evaluated intestinal permeability using the FITC-labeled dextran method and analysed tight junction proteins expression by immunofluorescence and PCR. We also measured CD4⁺FoxP3⁺ regulatory T cells proportion by FACS analysis, microbiota composition by pyrosequencing, and local cytokine production by ELISA. Lc leads to a significant protection against increased intestinal permeability and barrier dysfunction shown by preserved ZO-1 expression. We found that the Lc treatment increases the numbers of CD4⁺FoxP3⁺ regulatory T cells in mesenteric lymph nodes (MLN), decreases production of pro-inflammatory cytokines TNF-α and IFN-γ, and anti-inflammatory IL-10 in Peyer''s patches and large intestine, and changes the gut microbiota composition. Moreover, Lc treatment prevents lipopolysaccharide-induced TNF-α expression in RAW 264.7 cell line by down-regulating the NF-κB signaling pathway.

Conclusion/Significance

Our study provided evidence that even non-living probiotic bacteria can prevent the development of severe forms of intestinal inflammation by strengthening the integrity of intestinal barrier and modulation of gut microenvironment.     

Imaging spectroscopy reveals the effects of topography and logging on the leaf chemistry of tropical forest canopy trees

Logging, pervasive across the lowland tropics, affects millions of hectares of forest, yet its influence on nutrient cycling remains poorly understood. One hypothesis is that logging influences phosphorus (P) cycling, because this scarce nutrient is removed in extracted timber and eroded soil, leading to shifts in ecosystem functioning and community composition. However, testing this is challenging because P varies within landscapes as a function of geology, topography and climate. Superimposed upon these trends are compositional changes in logged forests, with species with more acquisitive traits, characterized by higher foliar P concentrations, more dominant. It is difficult to resolve these patterns using traditional field approaches alone. Here, we use airborne light detection and ranging‐guided hyperspectral imagery to map foliar nutrient (i.e. P, nitrogen [N]) concentrations, calibrated using field measured traits, over 400 km² of northeastern Borneo, including a landscape‐level disturbance gradient spanning old‐growth to repeatedly logged forests. The maps reveal that canopy foliar P and N concentrations decrease with elevation. These relationships were not identified using traditional field measurements of leaf and soil nutrients. After controlling for topography, canopy foliar nutrient concentrations were lower in logged forest than in old‐growth areas, reflecting decreased nutrient availability. However, foliar nutrient concentrations and specific leaf area were greatest in relatively short patches in logged areas, reflecting a shift in composition to pioneer species with acquisitive traits. N:P ratio increased in logged forest, suggesting reduced soil P availability through disturbance. Through the first landscape scale assessment of how functional leaf traits change in response to logging, we find that differences from old‐growth forest become more pronounced as logged forests increase in stature over time, suggesting exacerbated phosphorus limitation as forests recover.     

Investigating the relationship between climate,stand age,and temporal trends in masting behavior of European forest trees

Masting—temporally variable seed production with high spatial synchrony—is a pervasive strategy in wind‐pollinated trees that is hypothesized to be vulnerable to climate change due to its correlation with variability in abiotic conditions. Recent work suggests that aging may also have strong effects on seed production patterns of trees, but this potential confounding factor has not been considered in previous times series analysis of climate change effects. Using a 54 year dataset for seven dominant species in 17 forests across Poland, we used the proportion of seed‐producing trees (PST) to contrast the predictions of the climate change and aging hypotheses in Abies alba, Fagus sylvatica, Larix decidua, Picea abies, Pinus sylvestris, Quercus petraea, and Quercus robur. Our results show that in all species, PST increased over time and that this change correlated most strongly with stand age, while the standardized precipitation–evapotranspiration index, a measure of drought, contributed to temporal trends in PST of F. sylvatica and Q. robur. Temporal variability of PST also increased over time in all species except P. sylvestris, while trends in temporal autocorrelation and among‐stand synchrony reflect species‐specific masting strategies. Our results suggest a pivotal role of plant ontogeny in driving not only the extent but also variability and synchrony of reproduction in temperate forest trees. In a time of increasing forest regrowth in Europe, we therefore call for increased attention to demographic effects such as aging on plant reproductive behavior, particularly in studies examining global change effects using long‐term time series data.     

Autumn-winter diet overlap of fallow, red, and roe deer in forest ecosystems, Southern Poland

The wild population of fallow deer in Central Europe has grown considerably over the last decade. However, information on feeding habits of this alien species in relation to the indigenous red deer or roe deer, in areas of their co-occurrence, is scarce. A prevailing view maintains that their food-niches are distinct, although direct comparative studies have not been carried out. Therefore, the aim of the research was to compare the diets of fallow, red, and roe deer feeding in the same habitat. Research was based on the rumen contents of 242 animals hunted in the autumn-winter season in the forests of Southern Poland. The analyses demonstrated that fallow deer are moderate grazers in such conditions and eat more graminoids in comparison to red or roe deer (36.4% vs. 16.1% or 5.5%, respectively). On the other hand, it feeds on less browse (17.2% vs. 41.4%) or dwarf shrubs (8.4% vs. 19.0%) than red deer, and on less bramble (10.9% vs. 34.6%) or forbs (4.0% vs. 7.6%) in comparison to roe deer (P=0.05). Although the diets of the three deer species differ in terms of the proportion of each food type in their diet, overlapping of their food-niches is high (52.6%).