Expression of human cathepsin L or human cathepsin V in mouse thymus mediates positive selection of T helper cells in cathepsin L knock-out mice

A genetic deficiency of the cysteine protease cathepsin L (Ctsl) in mice results in impaired positive selection of conventional CD4+ T helper cells as a result of an incomplete processing of the MHC class II associated invariant chain or incomplete proteolytic generation of positively selecting peptide ligands. The human genome encodes, in contrast to the mouse genome, for two cathepsin L proteases, namely cathepsin L (CTSL) and cathepsin V (CTSV; alternatively cathepsin L2). In the human thymic cortex, CTSV is the predominately expressed protease as compared to CTSL or other cysteine cathepsins. In order to analyze the functions of CTSL and CTSV in the positive selection of CD4+ T cells we employed Ctsl knock-out mice crossed either with transgenic mice expressing CTSL under the control of its genuine human promoter or with transgenic mice expressing CTSV under the control of the keratin 14 (K14) promoter, which drives expression to the cortical epithelium. Both human proteases are expressed in the thymus of the transgenic mice, and independent expression of both CTSL and CTSV rescues the reduced frequency of CD4+ T cells in Ctsl-deficient mice. Moreover, the expression of the human cathepsins does not change the number of CD4+CD25+Foxp3+ regulatory T cells, but the normalization of the frequency of conventional CD4+ T cell in the transgenic mice results in a rebalancing of conventional T cells and regulatory T cells. We conclude that the functional differences of CTSL and CTSV in vivo are not mainly determined by their inherent biochemical properties, but rather by their tissue specific expression pattern.

Serodiagnosis of Echinococcus spp. infection: explorative selection of diagnostic antigens by peptide microarray

BACKGROUND: Production of native antigens for serodiagnosis of helminthic infections is laborious and hampered by batch-to-batch variation. For serodiagnosis of echinococcosis, especially cystic disease, most screening tests rely on crude or purified Echinococcus granulosus hydatid cyst fluid. To resolve limitations associated with native antigens in serological tests, the use of standardized and highly pure antigens produced by chemical synthesis offers considerable advantages, provided appropriate diagnostic sensitivity and specificity is achieved. METHODOLOGY/PRINCIPAL FINDINGS: Making use of the growing collection of genomic and proteomic data, we applied a set of bioinformatic selection criteria to a collection of protein sequences including conceptually translated nucleotide sequence data of two related tapeworms, Echinococcus multilocularis and Echinococcus granulosus. Our approach targeted alpha-helical coiled-coils and intrinsically unstructured regions of parasite proteins potentially exposed to the host immune system. From 6 proteins of E. multilocularis and 5 proteins of E. granulosus, 45 peptides between 24 and 30 amino acids in length were designed. These peptides were chemically synthesized, spotted on microarrays and screened for reactivity with sera from infected humans. Peptides reacting above the cut-off were validated in enzyme-linked immunosorbent assays (ELISA). Peptides identified failed to differentiate between E. multilocularis and E. granulosus infection. The peptide performing best reached 57% sensitivity and 94% specificity. This candidate derived from Echinococcus multilocularis antigen B8/1 and showed strong reactivity to sera from patients infected either with E. multilocularis or E. granulosus. CONCLUSIONS/SIGNIFICANCE: This study provides proof of principle for the discovery of diagnostically relevant peptides by bioinformatic selection complemented with screening on a high-throughput microarray platform. Our data showed that a single peptide cannot provide sufficient diagnostic sensitivity whereas pooling several peptide antigens improved sensitivity; thus combinations of several peptides may lead the way to new diagnostic tests that replace, or at least complement conventional immunodiagnosis of echinococcosis. Our strategy could prove useful for diagnostic developments in other pathogens.

Defining new criteria for selection of cell-based intestinal models using publicly available databases

The criteria for choosing relevant cell lines among a vast panel of available intestinal-derived lines exhibiting a wide range of functional properties are still ill-defined. The objective of this study was, therefore, to establish objective criteria for choosing relevant cell lines to assess their appropriateness as tumor models as well as for drug absorption studies.

Effect of Methodological and Ecological Approaches on Heterogeneity of Nest-Site Selection of a Long-Lived Vulture

The application of scientific-based conservation measures requires that sampling methodologies in studies modelling similar ecological aspects produce comparable results making easier their interpretation. We aimed to show how the choice of different methodological and ecological approaches can affect conclusions in nest-site selection studies along different Palearctic meta-populations of an indicator species. First, a multivariate analysis of the variables affecting nest-site selection in a breeding colony of cinereous vulture (Aegypius monachus) in central Spain was performed. Then, a meta-analysis was applied to establish how methodological and habitat-type factors determine differences and similarities in the results obtained by previous studies that have modelled the forest breeding habitat of the species. Our results revealed patterns in nesting-habitat modelling by the cinereous vulture throughout its whole range: steep and south-facing slopes, great cover of large trees and distance to human activities were generally selected. The ratio and situation of the studied plots (nests/random), the use of plots vs. polygons as sampling units and the number of years of data set determined the variability explained by the model. Moreover, a greater size of the breeding colony implied that ecological and geomorphological variables at landscape level were more influential. Additionally, human activities affected in greater proportion to colonies situated in Mediterranean forests. For the first time, a meta-analysis regarding the factors determining nest-site selection heterogeneity for a single species at broad scale was achieved. It is essential to homogenize and coordinate experimental design in modelling the selection of species' ecological requirements in order to avoid that differences in results among studies would be due to methodological heterogeneity. This would optimize best conservation and management practices for habitats and species in a global context.

Eluding oblivion with smart stochastic selection of synaptic updates

The variables involved in the equations that describe realistic synaptic dynamics always vary in a limited range. Their boundedness makes the synapses forgetful, not for the mere passage of time, but because new experiences overwrite old memories. The forgetting rate depends on how many synapses are modified by each new experience: many changes means fast learning and fast forgetting, whereas few changes means slow learning and long memory retention. Reducing the average number of modified synapses can extend the memory span at the price of a reduced amount of information stored when a new experience is memorized. Every trick which allows to slow down the learning process in a smart way can improve the memory performance. We review some of the tricks that allow to elude fast forgetting (oblivion). They are based on the stochastic selection of the synapses whose modifications are actually consolidated following each new experience. In practice only a randomly selected, small fraction of the synapses eligible for an update are actually modified. This allows to acquire the amount of information necessary to retrieve the memory without compromising the retention of old experiences. The fraction of modified synapses can be further reduced in a smart way by changing synapses only when it is really necessary, i.e. when the post-synaptic neuron does not respond as desired. Finally we show that such a stochastic selection emerges naturally from spike driven synaptic dynamics which read noisy pre and post-synaptic neural activities. These activities can actually be generated by a chaotic system.

Selection of preadapted populations allowed Senecio inaequidens to invade Central Europe

Invasive species often evolve rapidly in response to the novel biotic and abiotic conditions in their introduced range. Such adaptive evolutionary changes might play an important role in the success of some invasive species. Here, we investigated whether introduced European populations of the South African ragwort Senecio inaequidens (Asteraceae) have genetically diverged from native populations. We carried out a greenhouse experiment where 12 South African and 11 European populations were for several months grown at two levels of nutrient availability, as well as in the presence or absence of a generalist insect herbivore. We found that, in contrast to a current hypothesis, plants from introduced populations had a significantly lower reproductive output, but higher allocation to root biomass, and they were more tolerant to insect herbivory. Moreover, introduced populations were less genetically variable, but displayed greater plasticity in response to fertilization. Finally, introduced populations were phenotypically most similar to a subset of native populations from mountainous regions in southern Africa. Taking into account the species' likely history of introduction, our data support the idea that the invasion success of Senecio inaequidens in Central Europe is based on selective introduction of specific preadapted and plastic genotypes rather than on adaptive evolution in the introduced range.