Genetic variants in toll-like receptors are not associated with rheumatoid arthritis susceptibility or anti-tumour necrosis factor treatment outcome

Background

Several studies point to a role of Toll-like receptors (TLRs) in the development of rheumatoid arthritis (RA). We investigated if genetic variants in TLR genes are associated with RA and response to tumour necrosis factor blocking (anti-TNF) medication.

Methodology and Principal Findings

22 single nucleotide polymorphisms (SNPs) in seven TLR genes were genotyped in a Dutch cohort consisting of 378 RA patients and 294 controls. Significantly associated variants were investigated in replication cohorts from The Netherlands, United Kingdom and Sweden (2877 RA patients and 2025 controls). 182 of the Dutch patients were treated with anti-TNF medication. Using these patients and a replication cohort (269 Swedish patients) we analysed if genetic variants in TLR genes were associated with anti-TNF outcome. In the discovery phase of the study we found a significant association of SNPs rs2072493 in TLR5 and rs3853839 in TLR7 with RA disease susceptibility. Meta-analysis of discovery and replication cohorts did not confirm these findings. SNP rs2072493 in TLR5 was associated with anti-TNF outcome in the Dutch but not in the Swedish population.

Conclusion

We conclude that genetic variants in TLRs do not play a major role in susceptibility for developing RA nor in anti-TNF treatment outcome in a Caucasian population.     

The energetics of Calanus euxinus: locomotion, filtration of food and specific dynamic action

The effect of locomotor activity on respiration rate was studiedin the food-deprived copepod Calanus euxinus tethered to a forcesensor. The power generated by mouth appendages during cruisinglocomotion, with a frequency of 40 Hz, accounted for 0.026 and0.0031 W for metabolic and mechanical processes, respectively.To overcome total hydrodynamic drag during foraging with a meanswimming speed of 3.2 cm s^–1, the copepods need 0.4 x10^–3 W, equating to 1.3% of total metabolism. The lossesof mechanical energy for body propulsion amounted to 1.3 x 10^–3W, whilst the cost of feeding current generation run up to 1.8x 10^–3 W, or 58% of the total. Changing of locomotor activityand respiration rate during feeding was examined separatelyin tethered and free-swimming copepods. At algal concentrationof 300 µg C L^–1, the magnitude of specific dynamicaction (SDA) averaged 1.2 ± 0.44 nL O₂ µg C^–1h^–1 in copepodites V and females, with similar movingactivity before and during feeding. The contribution of SDAinto total metabolism varied from 23 to 85% in C. euxinus withlow activity level and constituted only 10% in active animals.In starved copepods, with low locomotor activity, feeding eventsstimulated the increase in frequency and total duration of locomotionwhich resulted in elevated energy expenditure enhancing the‘apparent SDA’.     

Analysis of transduction in wastewater bacterial populations by targeting the phage-derived 16S rRNA gene sequences

Bacterial 16S rRNA genes transduced by bacteriophages were identified and analyzed in order to estimate the extent of the bacteriophage-mediated horizontal gene transfer in the wastewater environment. For this purpose, phage and bacterial DNA was isolated from the oxidation tank of a municipal wastewater treatment plant. Phylogenetic analysis of the 16S rRNA gene sequences cloned from a phage metagenome revealed that bacteriophages transduce genetic material in several major groups of bacteria. The groups identified were as follows: Betaproteobacteria, Gammaproteobacteria, Alphaproteobacteria, Actinomycetales and Firmicutes. Analysis of the 16S rRNA gene sequences in the total bacterial DNA from the same sample revealed that several bacterial groups found in the oxidation tank were not present in the phage metagenome (e.g. Deltaproteobacteria, Nitrospira, Planctomycetes and many Actinobacteria genera). These results suggest that transduction in a wastewater environment occurs in several bacterial groups; however, not all species are equally involved into this process. The data also showed that a number of distinctive bacterial strains participate in transduction-mediated gene transfer within identified bacterial groupings. Denaturing gradient gel electrophoresis analysis confirmed that profiles of the transduced 16S rRNA gene sequences and those present in the whole microbial community show significant differences.     

Spin Trapping Using 2,2-Dimethyl-2H-Imidazole-1-Oxides

The ability of novel cyclic nitrones, 4-substituted 2,2-dimethyl-2H-imidazole-1-oxides (IMO's) to trap a variety of short-lived free radicals has been investigated using ESR spectroscopy. IMO's scavenge oxygen-, carbon- and sulfur-derived free radicals to give persistent nitroxides. Compared to the spin trap 5,5-dimethyl-pyrroline-1-oxide, a higher lifetime of hydroxyl radical adducts and a higher selectivity related to the trapping of carbon-centered radicals was found. A reaction between IMO's and superoxide was not observed. ESR parameters of 4-carboxyl-2,2-dimethyl-2H-imidazole-1-oxide (CIMO) spin adducts are highly sensitive to the structure of the trapped radical, e.g., different spectra were detected with radicals derived from Na₂SO₃ and NaHSO₃. From the data obtained, a successful application of these new spin traps in biological systems can be expected.     

Ditches as species‐rich secondary habitats and refuge for meadow species in agricultural marsh grasslands

Questions

Can drainage ditches in agricultural marsh grassland provide a suitable habitat for the persistence of fen meadow species? How does the ditch margin vegetation develop as a function of regular dredging? Is ornithologically oriented management also beneficial for plant biodiversity?

Location

Riparian marshes, Eider‐Treene‐Sorge lowland, Schleswig‐Holstein, Germany.

Methods

We performed vegetation surveys of drainage ditches along with their water body, slope and margin structures annually for 3 years. The data were analysed with respect to date and means of ditch dredging. In addition, we recorded vegetation of the surrounding agricultural grassland, measured nutrient status of the soil and the water body and sampled seed bank of the ditch slopes. We used ANOVA and multivariate methods to describe the development of the ditch vegetation and the persistence of target meadow species.

Results

Vegetation re‐development of ditch margins proceeds quite rapidly after disturbance from dredging. Dominance of mudbank species was observed only in the first year, followed by an increase of reed species and reduction of phytodiversity. Target species of wet meadow communities reach highest abundance in the second and third year and build a significant seed bank before being suppressed by reeds.

Conclusions

In heavily eutrophicated, intensively used marsh grassland, regularly disturbed ditch margins are important secondary habitats for pioneer and subdominant wetland species, which have nearly disappeared in a larger area. Current management cycles of ditch dredging every 3–4 years comply with the successional development, allowing the mudbank and wet meadow species to persist in the vegetation and seed bank. In contrast to the frequency, the form of dredging (ditch profile), which is crucial for bird protection, plays a minor role for plants. We recommend moderate disturbance (mowing of ditch margins) to suppress strong competitors in the years between dredging for additional support to target plant species.     

Extensive metabolic cross-talk in melon fruit revealed by spatial and developmental combinatorial metabolomics

? Variations in tissue development and spatial composition have a major impact on the nutritional and organoleptic qualities of ripe fleshy fruit, including melon (Cucumis melo). To gain a deeper insight into the mechanisms involved in these changes, we identified key metabolites for rational food quality design. ? The metabolome, volatiles and mineral elements were profiled employing an unprecedented range of complementary analytical technologies. Fruits were followed at a number of time points during the final ripening process and tissues were collected across the fruit flesh from rind to seed cavity. Approximately 2000 metabolite signatures and 15 mineral elements were determined in an assessment of temporal and spatial melon fruit development. ? This study design enabled the identification of: coregulated hubs (including aspartic acid, 2-isopropylmalic acid, β-carotene, phytoene and dihydropseudoionone) in metabolic association networks; global patterns of coordinated compositional changes; and links of primary and secondary metabolism to key mineral and volatile fruit complements. ? The results reveal the extent of metabolic interactions relevant to ripe fruit quality and thus have enabled the identification of essential candidate metabolites for the high-throughput screening of melon breeding populations for targeted breeding programmes aimed at nutrition and flavour improvement.     

Expression Quantitative Trait Loci Are Highly Sensitive to Cellular Differentiation State

Genetical genomics is a strategy for mapping gene expression variation to expression quantitative trait loci (eQTLs). We performed a genetical genomics experiment in four functionally distinct but developmentally closely related hematopoietic cell populations isolated from the BXD panel of recombinant inbred mouse strains. This analysis allowed us to analyze eQTL robustness/sensitivity across different cellular differentiation states. Although we identified a large number (365) of “static” eQTLs that were consistently active in all four cell types, we found a much larger number (1,283) of “dynamic” eQTLs showing cell-type–dependence. Of these, 140, 45, 531, and 295 were preferentially active in stem, progenitor, erythroid, and myeloid cells, respectively. A detailed investigation of those dynamic eQTLs showed that in many cases the eQTL specificity was associated with expression changes in the target gene. We found no evidence for target genes that were regulated by distinct eQTLs in different cell types, suggesting that large-scale changes within functional regulatory networks are uncommon. Our results demonstrate that heritable differences in gene expression are highly sensitive to the developmental stage of the cell population under study. Therefore, future genetical genomics studies should aim at studying multiple well-defined and highly purified cell types in order to construct as comprehensive a picture of the changing functional regulatory relationships as possible.     

Gene-gene and gene-environment interactions involving HLA-DRB1, PTPN22, and smoking in two subsets of rheumatoid arthritis

Gene-gene and gene-environment interactions are key features in the development of rheumatoid arthritis (RA) and other complex diseases. The aim of this study was to use and compare three different definitions of interaction between the two major genetic risk factors of RA--the HLA-DRB1 shared epitope (SE) alleles and the PTPN22 R620W allele--in three large case-control studies: the Swedish Epidemiological Investigation of Rheumatoid Arthritis (EIRA) study, the North American RA Consortium (NARAC) study, and the Dutch Leiden Early Arthritis Clinic study (in total, 1,977 cases and 2,405 controls). The EIRA study was also used to analyze interactions between smoking and the two genes. "Interaction" was defined either as a departure from additivity, as interaction in a multiplicative model, or in terms of linkage disequilibrium--for example, deviation from independence of penetrance of two unlinked loci. Consistent interaction, defined as departure from additivity, between HLA-DRB1 SE alleles and the A allele of PTPN22 R620W was seen in all three studies regarding anti-CCP-positive RA. Testing for multiplicative interactions demonstrated an interaction between the two genes only when the three studies were pooled. The linkage disequilibrium approach indicated a gene-gene interaction in EIRA and NARAC, as well as in the pooled analysis. No interaction was seen between smoking and PTPN22 R620W. A new pattern of interactions is described between the two major known genetic risk factors and the major environmental risk factor concerning the risk of developing anti-CCP-positive RA. The data extend the basis for a pathogenetic hypothesis for RA involving genetic and environmental factors. The study also raises and illustrates principal questions concerning ways to define interactions in complex diseases.     

Species introduction in restoration projects – Evaluation of different techniques for the establishment of semi-natural grasslands in Central and Northwestern Europe

During recent decades, many studies have shown that the successful restoration of species-rich grasslands is often seed-limited because of depleted seed banks and limited seed dispersal in modern fragmented landscapes. In Europe, commercial seed mixtures, which are widely used for restoration measures, mostly consist of species and varieties of non-local provenance. The regional biodiversity of a given landscape, however, can be preserved only when seeds or plants of local provenance are used in restoration projects. Furthermore, the transfer of suitable target species of local provenance can strongly enhance restoration success.We review and evaluate the success of currently used near-natural methods for the introduction of target plant species (e.g. seeding of site-specific seed mixtures, transfer of fresh seed-containing hay, vacuum harvesting, transfer of turves or seed-containing soil) on restoration sites, ranging from dry and mesic meadows to floodplain grasslands and fens. Own data combined with literature findings show species establishment rates during the initial phase as well as the persistence of target species during long-term vegetation development on restoration sites.In conclusion, our review indicates that seed limitation can be overcome successfully by most of the reviewed measures for species introduction. The establishment of species-rich grasslands is most successful when seeds, seed-containing plant material or soil are spread on bare soil of ex-arable fields after tilling or topsoil removal, or on raw soils, e.g. in mined areas. In species-poor grasslands without soil disturbance and on older ex-arable fields with dense weed vegetation, final transfer rates were the lowest. For future restoration projects, suitable measures have to be chosen carefully from case to case as they differ considerably in costs and logistic effort. Long-term prospects for restored grassland are especially good when management can be incorporated in agricultural systems.     

Class II fusion protein of alphaviruses drives membrane fusion through the same pathway as class I proteins

Viral fusion proteins of classes I and II differ radically in their initial structures but refold toward similar conformations upon activation. Do fusion pathways mediated by alphavirus E1 and influenza virus hemagglutinin (HA) that exemplify classes II and I differ to reflect the difference in their initial conformations, or concur to reflect the similarity in the final conformations? Here, we dissected the pathway of low pH–triggered E1-mediated cell–cell fusion by reducing the numbers of activated E1 proteins and by blocking different fusion stages with specific inhibitors. The discovered progression from transient hemifusion to small, and then expanding, fusion pores upon an increase in the number of activated fusion proteins parallels that established for HA-mediated fusion. We conclude that proteins as different as E1 and HA drive fusion through strikingly similar membrane intermediates, with the most energy-intensive stages following rather than preceding hemifusion. We propose that fusion reactions catalyzed by all proteins of both classes follow a similar pathway.