Novel prokaryotic expression of thioredoxin-fused insulinoma associated protein tyrosine phosphatase 2 (IA-2), its characterization and immunodiagnostic application

Background The insulinoma associated protein tyrosine phosphatase 2 (IA-2) is one of the immunodominant autoantigens involved in the autoimmune attack to the beta-cell in Type 1 Diabetes Mellitus. In this work we have developed a complete and original process for the production and recovery of the properly folded intracellular domain of IA-2 fused to thioredoxin (TrxIA-2ic) in Escherichia coli GI698 and GI724 strains. We have also carried out the biochemical and immunochemical characterization of TrxIA-2icand design variants of non-radiometric immunoassays for the efficient detection of IA-2 autoantibodies (IA-2A). Results The main findings can be summarized in the following statements: i) TrxIA-2ic expression after 3 h of induction on GI724 strain yielded ≈ 10 mg of highly pure TrxIA-2ic/L of culture medium by a single step purification by affinity chromatography, ii) the molecular weight of TrxIA-2ic (55,358 Da) could be estimated by SDS-PAGE, size exclusion chromatography and mass spectrometry, iii) TrxIA-2ic was properly identified by western blot and mass spectrometric analysis of proteolytic digestions (63.25 % total coverage), iv) excellent immunochemical behavior of properly folded full TrxIA-2ic was legitimized by inhibition or displacement of [35S]IA-2 binding from IA-2A present in Argentinian Type 1 Diabetic patients, v) great stability over time was found under proper storage conditions and vi) low cost and environmentally harmless ELISA methods for IA-2A assessment were developed, with colorimetric or chemiluminescent detection. Conclusions E. coli GI724 strain emerged as a handy source of recombinant IA-2ic, achieving high levels of expression as a thioredoxin fusion protein, adequately validated and applicable to the development of innovative and cost-effective immunoassays for IA-2A detection in most laboratories.Fil: Guerra, Luciano Lucas. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Faccinetti, Natalia Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Trabucchi, Aldana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Rovitto, Bruno David. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Sabljic, Adriana Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Poskus, Edgardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Iacono, Ruben Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; ArgentinaFil: Valdez, Silvina Noemi. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Estudios de la Inmunidad Humoral Prof. Ricardo A. Margni; Argentin

Biological response of an in vitro human 3D lung cell model exposed to brake wear debris varies based on brake pad formulation

Wear particles from automotive friction brake pads of various sizes, morphology, and chemical composition are significant contributors towards particulate matter. Knowledge concerning the potential adverse effects following inhalation exposure to brake wear debris is limited. Our aim was, therefore, to generate brake wear particles released from commercial low-metallic and non-asbestos organic automotive brake pads used in mid-size passenger cars by a full-scale brake dynamometer with an environmental chamber simulating urban driving and to deduce their potential hazard in vitro. The collected fractions were analysed using scanning electron microscopy via energy-dispersive X-ray spectroscopy (SEM-EDS) and Raman microspectroscopy. The biological impact of the samples was investigated using a human 3D multicellular model consisting of human epithelial cells (A549) and human primary immune cells (macrophages and dendritic cells) mimicking the human epithelial tissue barrier. The viability, morphology, oxidative stress, and (pro-)inflammatory response of the cells were assessed following 24 h exposure to similar to 12, similar to 24, and similar to 48 A mu g/cm(2) of non-airborne samples and to similar to 3.7 A mu g/cm(2) of different brake wear size fractions (2-4, 1-2, and 0.25-1 A mu m) applying a pseudo-air-liquid interface approach. Brake wear debris with low-metallic formula does not induce any adverse biological effects to the in vitro lung multicellular model. Brake wear particles from non-asbestos organic formulated pads, however, induced increased (pro-)inflammatory mediator release from the same in vitro system. The latter finding can be attributed to the different particle compositions, specifically the presence of anatase.Web of Science9272351233

Speciation and fate of trace metals in estuarine sediments under reduced and oxidized conditions, Seaplane Lagoon, Alameda Naval Air Station (USA)

We have identified important chemical reactions that control the fate of metal-contaminated estuarine sediments if they are left undisturbed (in situ) or if they are dredged. We combined information on the molecular bonding of metals in solids from X-ray absorption spectroscopy (XAS) with thermodynamic and kinetic driving forces obtained from dissolved metal concentrations to deduce the dominant reactions under reduced and oxidized conditions. We evaluated the in situ geochemistry of metals (cadmium, chromium, iron, lead, manganese and zinc) as a function of sediment depth (to 100 cm) from a 60 year record of contamination at the Alameda Naval Air Station, California. Results from XAS and thermodynamic modeling of porewaters show that cadmium and most of the zinc form stable sulfide phases, and that lead and chromium are associated with stable carbonate, phosphate, phyllosilicate, or oxide minerals. Therefore, there is minimal risk associated with the release of these trace metals from the deeper sediments contaminated prior to the Clean Water Act (1975) as long as reducing conditions are maintained. Increased concentrations of dissolved metals with depth were indicative of the formation of metal HS(- )complexes. The sediments also contain zinc, chromium, and manganese associated with detrital iron-rich phyllosilicates and/or oxides. These phases are recalcitrant at near-neutral pH and do not undergo reductive dissolution within the 60 year depositional history of sediments at this site. The fate of these metals during dredging was evaluated by comparing in situ geochemistry with that of sediments oxidized by seawater in laboratory experiments. Cadmium and zinc pose the greatest hazard from dredging because their sulfides were highly reactive in seawater. However, their dissolved concentrations under oxic conditions were limited eventually by sorption to or co-precipitation with an iron (oxy)hydroxide. About 50% of the reacted CdS and 80% of the reacted ZnS were bonded to an oxide-substrate at the end of the 90-day oxidation experiment. Lead and chromium pose a minimal hazard from dredging because they are bonded to relatively insoluble carbonate, phosphate, phyllosilicate, or oxide minerals that are stable in seawater. These results point out the specific chemical behavior of individual metals in estuarine sediments, and the need for direct confirmation of metal speciation in order to constrain predictive models that realistically assess the fate of metals in urban harbors and coastal sediments

The Psychological Science Accelerator's COVID-19 rapid-response dataset

In response to the COVID-19 pandemic, the Psychological Science Accelerator coordinated three large-scale psychological studies to examine the effects of loss-gain framing, cognitive reappraisals, and autonomy framing manipulations on behavioral intentions and affective measures. The data collected (April to October 2020) included specific measures for each experimental study, a general questionnaire examining health prevention behaviors and COVID-19 experience, geographical and cultural context characterization, and demographic information for each participant. Each participant started the study with the same general questions and then was randomized to complete either one longer experiment or two shorter experiments. Data were provided by 73,223 participants with varying completion rates. Participants completed the survey from 111 geopolitical regions in 44 unique languages/dialects. The anonymized dataset described here is provided in both raw and processed formats to facilitate re-use and further analyses. The dataset offers secondary analytic opportunities to explore coping, framing, and self-determination across a diverse, global sample obtained at the onset of the COVID-19 pandemic, which can be merged with other time-sampled or geographic data

The Psychological Science Accelerator’s COVID-19 rapid-response dataset

In response to the COVID-19 pandemic, the Psychological Science Accelerator coordinated three large-scale psychological studies to examine the effects of loss-gain framing, cognitive reappraisals, and autonomy framing manipulations on behavioral intentions and affective measures. The data collected (April to October 2020) included specific measures for each experimental study, a general questionnaire examining health prevention behaviors and COVID-19 experience, geographical and cultural context characterization, and demographic information for each participant. Each participant started the study with the same general questions and then was randomized to complete either one longer experiment or two shorter experiments. Data were provided by 73,223 participants with varying completion rates. Participants completed the survey from 111 geopolitical regions in 44 unique languages/dialects. The anonymized dataset described here is provided in both raw and processed formats to facilitate re-use and further analyses. The dataset offers secondary analytic opportunities to explore coping, framing, and self-determination across a diverse, global sample obtained at the onset of the COVID-19 pandemic, which can be merged with other time-sampled or geographic data

Lymnaea schirazensis, an Overlooked Snail Distorting Fascioliasis Data: Genotype, Phenotype, Ecology, Worldwide Spread, Susceptibility, Applicability

BACKGROUND: Lymnaeid snails transmit medical and veterinary important trematodiases, mainly fascioliasis. Vector specificity of fasciolid parasites defines disease distribution and characteristics. Different lymnaeid species appear linked to different transmission and epidemiological patterns. Pronounced susceptibility differences to absolute resistance have been described among lymnaeid populations. When assessing disease characteristics in different endemic areas, unexpected results were obtained in studies on lymnaeid susceptibility to Fasciola. We undertook studies to understand this disease transmission heterogeneity. METHODOLOGY/PRINCIPAL FINDINGS: A ten-year study in Iran, Egypt, Spain, the Dominican Republic, Mexico, Venezuela, Ecuador and Peru, demonstrated that such heterogeneity is not due to susceptibility differences, but to a hitherto overlooked cryptic species, Lymnaea schirazensis, confused with the main vector Galba truncatula and/or other Galba/Fossaria vectors. Nuclear rDNA and mtDNA sequences and phylogenetic reconstruction highlighted an old evolutionary divergence from other Galba/Fossaria species, and a low intraspecific variability suggesting a recent spread from one geographical source. Morphometry, anatomy and egg cluster analyses allowed for phenotypic differentiation. Selfing, egg laying, and habitat characteristics indicated a migration capacity by passive transport. Studies showed that it is not a vector species (n = 8572 field collected, 20 populations): snail finding and penetration by F. hepatica miracidium occur but never lead to cercarial production (n = 338 experimentally infected). CONCLUSIONS/SIGNIFICANCE: This species has been distorting fasciolid specificity/susceptibility and fascioliasis geographical distribution data. Hence, a large body of literature on G. truncatula should be revised. Its existence has henceforth to be considered in research. Genetic data on livestock, archeology and history along the 10,000-year post-domestication period explain its wide spread from the Neolithic Fertile Crescent. It is an efficient biomarker for the follow-up of livestock movements, a crucial aspect in fascioliasis emergence. It offers an outstanding laboratory model for genetic studies on susceptibility/resistance in F. hepatica/lymnaeid interaction, a field of applied research with disease control perspectives

Climatic controls of decomposition drive the global biogeography of forest-tree symbioses

The identity of the dominant root-associated microbial symbionts in a forest determines the ability of trees to access limiting nutrients from atmospheric or soil pools1,2, sequester carbon3,4 and withstand the effects of climate change5,6. Characterizing the global distribution of these symbioses and identifying the factors that control this distribution are thus integral to understanding the present and future functioning of forest ecosystems. Here we generate a spatially explicit global map of the symbiotic status of forests, using a database of over 1.1 million forest inventory plots that collectively contain over 28,000 tree species. Our analyses indicate that climate variables—in particular, climatically controlled variation in the rate of decomposition—are the primary drivers of the global distribution of major symbioses. We estimate that ectomycorrhizal trees, which represent only 2% of all plant species7, constitute approximately 60% of tree stems on Earth. Ectomycorrhizal symbiosis dominates forests in which seasonally cold and dry climates inhibit decomposition, and is the predominant form of symbiosis at high latitudes and elevation. By contrast, arbuscular mycorrhizal trees dominate in aseasonal, warm tropical forests, and occur with ectomycorrhizal trees in temperate biomes in which seasonally warm-and-wet climates enhance decomposition. Continental transitions between forests dominated by ectomycorrhizal or arbuscular mycorrhizal trees occur relatively abruptly along climate-driven decomposition gradients; these transitions are probably caused by positive feedback effects between plants and microorganisms. Symbiotic nitrogen fixers—which are insensitive to climatic controls on decomposition (compared with mycorrhizal fungi)—are most abundant in arid biomes with alkaline soils and high maximum temperatures. The climatically driven global symbiosis gradient that we document provides a spatially explicit quantitative understanding of microbial symbioses at the global scale, and demonstrates the critical role of microbial mutualisms in shaping the distribution of plant species

The global biogeography of tree leaf form and habit

Understanding what controls global leaf type variation in trees is crucial for comprehending their role in terrestrial ecosystems, including carbon, water and nutrient dynamics. Yet our understanding of the factors influencing forest leaf types remains incomplete, leaving us uncertain about the global proportions of needle-leaved, broadleaved, evergreen and deciduous trees. To address these gaps, we conducted a global, ground-sourced assessment of forest leaf-type variation by integrating forest inventory data with comprehensive leaf form (broadleaf vs needle-leaf) and habit (evergreen vs deciduous) records. We found that global variation in leaf habit is primarily driven by isothermality and soil characteristics, while leaf form is predominantly driven by temperature. Given these relationships, we estimate that 38% of global tree individuals are needle-leaved evergreen, 29% are broadleaved evergreen, 27% are broadleaved deciduous and 5% are needle-leaved deciduous. The aboveground biomass distribution among these tree types is approximately 21% (126.4 Gt), 54% (335.7 Gt), 22% (136.2 Gt) and 3% (18.7 Gt), respectively. We further project that, depending on future emissions pathways, 17-34% of forested areas will experience climate conditions by the end of the century that currently support a different forest type, highlighting the intensification of climatic stress on existing forests. By quantifying the distribution of tree leaf types and their corresponding biomass, and identifying regions where climate change will exert greatest pressure on current leaf types, our results can help improve predictions of future terrestrial ecosystem functioning and carbon cycling