pH-induced molecular shedding drives the formation of amyloid fibril-derived oligomers

Amyloid disorders cause debilitating illnesses through the formation of toxic protein aggregates. The mechanisms of amyloid toxicity and the nature of species responsible for mediating cellular dysfunction remain unclear. Here, using β2-microglobulin (β2m) as a model system, we show that the disruption of membranes by amyloid fibrils is caused by the molecular shedding of membrane-active oligomers in a process that is dependent on pH. Using thioflavin T (ThT) fluorescence, NMR, EM and fluorescence correlation spectroscopy (FCS), we show that fibril disassembly at pH 6.4 results in the formation of nonnative spherical oligomers that disrupt synthetic membranes. By contrast, fibril dissociation at pH 7.4 results in the formation of nontoxic, native monomers. Chemical cross-linking or interaction with hsp70 increases the kinetic stability of fibrils and decreases their capacity to cause membrane disruption and cellular dysfunction. The results demonstrate how pH can modulate the deleterious effects of preformed amyloid aggregates and suggest why endocytic trafficking through acidic compartments may be a key factor in amyloid disease

Self- and peer assessment may not be an accurate measure of PBL tutorial process

<p>Abstract</p> <p>Background</p> <p>Universidade Cidade de São Paulo adopted a problem-based learning (PBL) strategy as the predominant method for teaching and learning medicine. Self-, peer- and tutor marks of the educational process are taken into account as part of the final grade, which also includes assessment of content. This study compared the different perspectives (and grades) of evaluators during tutorials with first year medical students, from 2004 to 2007 (n = 349), from seven semesters.</p> <p>Methods</p> <p>The tutorial evaluation method was comprised of the students' self assessment (SA) (10%), tutor assessment (TA) (80%) and peer assessment (PA) (10%) to calculate a final educational process grade for each tutorial. We compared these three grades from each tutorial for seven semesters using ANOVA and a post hoc test.</p> <p>Results</p> <p>A total of 349 students participated with 199 (57%) women and 150 (42%) men. The SA and PA scores were consistently greater than the TA scores. Moreover, the SA and PA groups did not show statistical difference in any semester evaluated, while both differed from tutor assessment in all semesters (Kruskal-Wallis, Dunn's test). The Spearman rank order showed significant (p < 0.0001) and positive correlation for the SA and PA groups (r = 0.806); this was not observed when we compared TA with PA (r = 0.456) or TA with SA (r = 0.376).</p> <p>Conclusion</p> <p>Peer- and self-assessment marks might be reliable but not valid for PBL tutorial process, especially if these assessments are used for summative assessment, composing the final grade. This article suggests reconsideration of the use of summative assessment for self-evaluation in PBL tutorials.</p

Pathways for scale and discipline reconciliation: current socio-ecological modelling methodologies to explore and reconstitute human prehistoric dynamics

International audienceThis communication elaborates a plea for the necessity of a specific modelling methodology which does not sacrifice two modelling principles: explanation Micro and correlation Macro. Three goals are assigned to modelling strategies: describe, understand and predict. One tendency in historical and spatial modelling is to develop models at a micro level in order to describe and by that way, understand the connection between local ecological contexts, acquired through local ecological data, and local social practices, acquired through archaeology. However, such a method faces difficulties for expanding its validity: It is validated by its adequacy with local data, but the prediction step is unreachable and quite nothing can be said for places out where. On the other hand, building models at a far larger scale, for instance at the continent and even the world level, enhances the connection between ecology and its temporal variability. Such connections are based on well-founded theories but lower the " small causes, big effects " emergence corresponding to agent-based approaches and the related inherent variability of socio-ecological dynamics that one can notice at a lower scale. We then propose a plea for combining both elements for building large-scale modelling tools, which aims are to describe and provide predictions on long-term past evolutions, that include the test of explaining socio-anthropological hypotheses, i.e. the emergence and the spread of local social innovations

Transthyretin Aggregation Pathway toward the Formation of Distinct Cytotoxic Oligomers

Characterization of small oligomers formed at an early stage of amyloid formation is critical to understanding molecular mechanism of pathogenic aggregation process. Here we identifed and characterized cytotoxic oligomeric intermediates populated during transthyretin (TTR) aggregation process. Under the amyloid-forming conditions, TTR initially forms a dimer through interactions between outer strands. The dimers are then associated to form a hexamer with a spherical shape, which serves as a building block to self-assemble into cytotoxic oligomers. Notably, wild-type (WT) TTR tends to form linear oligomers, while aTTR variant(G53A) prefers forming annular oligomers with pore-like structures. Structural analyses of the amyloidogenic intermediates using circular dichroism (CD) and solid-state NMR revealthatthe dimer and oligomers have a signifcant degree of native-like β-sheet structures (35–38%), but with more disordered regions (~60%)than those of nativeTTR.TheTTR variant oligomers are also less structured than WT oligomers. The partially folded nature of the oligomeric intermediates might be a common structural property of cytotoxic oligomers.The higher fexibility of the dimer and oligomers may also compensate for the entropic loss due to the oligomerization of the monomers

Life expectancy associated with different ages at diagnosis of type 2 diabetes in high-income countries: 23 million person-years of observation

Background: The prevalence of type 2 diabetes is increasing rapidly, particularly among younger age groups. Estimates suggest that people with diabetes die, on average, 6 years earlier than people without diabetes. We aimed to provide reliable estimates of the associations between age at diagnosis of diabetes and all-cause mortality, cause-specific mortality, and reductions in life expectancy. Methods: For this observational study, we conducted a combined analysis of individual-participant data from 19 high-income countries using two large-scale data sources: the Emerging Risk Factors Collaboration (96 cohorts, median baseline years 1961–2007, median latest follow-up years 1980–2013) and the UK Biobank (median baseline year 2006, median latest follow-up year 2020). We calculated age-adjusted and sex-adjusted hazard ratios (HRs) for all-cause mortality according to age at diagnosis of diabetes using data from 1 515 718 participants, in whom deaths were recorded during 23·1 million person-years of follow-up. We estimated cumulative survival by applying age-specific HRs to age-specific death rates from 2015 for the USA and the EU. Findings: For participants with diabetes, we observed a linear dose–response association between earlier age at diagnosis and higher risk of all-cause mortality compared with participants without diabetes. HRs were 2·69 (95% CI 2·43–2·97) when diagnosed at 30–39 years, 2·26 (2·08–2·45) at 40–49 years, 1·84 (1·72–1·97) at 50–59 years, 1·57 (1·47–1·67) at 60–69 years, and 1·39 (1·29–1·51) at 70 years and older. HRs per decade of earlier diagnosis were similar for men and women. Using death rates from the USA, a 50-year-old individual with diabetes died on average 14 years earlier when diagnosed aged 30 years, 10 years earlier when diagnosed aged 40 years, or 6 years earlier when diagnosed aged 50 years than an individual without diabetes. Using EU death rates, the corresponding estimates were 13, 9, or 5 years earlier. Interpretation: Every decade of earlier diagnosis of diabetes was associated with about 3–4 years of lower life expectancy, highlighting the need to develop and implement interventions that prevent or delay the onset of diabetes and to intensify the treatment of risk factors among young adults diagnosed with diabetes. Funding: British Heart Foundation, Medical Research Council, National Institute for Health and Care Research, and Health Data Research UK

Outer membrane protein folding from an energy landscape perspective

The cell envelope is essential for the survival of Gram-negative bacteria. This specialised membrane is densely packed with outer membrane proteins (OMPs), which perform a variety of functions. How OMPs fold into this crowded environment remains an open question. Here, we review current knowledge about OFMP folding mechanisms in vitro and discuss how the need to fold to a stable native state has shaped their folding energy landscapes. We also highlight the role of chaperones and the β-barrel assembly machinery (BAM) in assisting OMP folding in vivo and discuss proposed mechanisms by which this fascinating machinery may catalyse OMP folding

An in vivo platform for identifying inhibitors of protein aggregation

Protein aggregation underlies an array of human diseases, yet only one small molecule therapeutic has been successfully developed to date. Here, we introduce an in vivo system, based on a β-lactamase tripartite fusion construct, capable of identifying aggregation-prone sequences in the periplasm of Escherichia coli and inhibitors that prevent their aberrant self-assembly. We demonstrate the power of the system using a range of proteins, from small unstructured peptides (islet amyloid polypeptide and amyloid β) to larger, folded immunoglobulin domains. Configured in a 48-well format, the split β-lactamase sensor readily differentiates between aggregation-prone and soluble sequences. Performing the assay in the presence of 109 compounds enabled a rank ordering of inhibition and revealed a new inhibitor of IAPP aggregation. This platform can be applied to both amyloidogenic and other aggregation-prone systems, independent of sequence or size, and can identify small molecules or other factors able to ameliorate or inhibit protein aggregation

A new era for understanding amyloid structures and disease

The aggregation of proteins into amyloid fibrils and their deposition into plaques and intracellular inclusions is the hallmark of amyloid disease. The accumulation and deposition of amyloid fibrils, collectively known as amyloidosis, is associated with many pathological conditions that can be associated with ageing, such as Alzheimer disease, Parkinson disease, type II diabetes and dialysis-related amyloidosis. However, elucidation of the atomic structure of amyloid fibrils formed from their intact protein precursors and how fibril formation relates to disease has remained elusive. Recent advances in structural biology techniques, including cryo-electron microscopy and solid-state NMR spectroscopy, have finally broken this impasse. The first near-atomic-resolution structures of amyloid fibrils formed in vitro, seeded from plaque material and analysed directly ex vivo are now available. The results reveal cross-β structures that are far more intricate than anticipated. Here, we describe these structures, highlighting their similarities and differences, and the basis for their toxicity. We discuss how amyloid structure may affect the ability of fibrils to spread to different sites in the cell and between organisms in a prion-like manner, along with their roles in disease. These molecular insights will aid in understanding the development and spread of amyloid diseases and are inspiring new strategies for therapeutic intervention

Harmful Elements in Estuarine and Coastal Systems

Estuaries and coastal zones are dynamic transitional systems which provide many economic and ecological benefits to humans, but also are an ideal habitat for other organisms as well. These areas are becoming contaminated by various anthropogenic activities due to a quick economic growth and urbanization. This chapter explores the sources, chemical speciation, sediment accumulation and removal mechanisms of the harmful elements in estuarine and coastal seawaters. It also describes the effects of toxic elements on aquatic flora and fauna. Finally, the toxic element pollution of the Venice Lagoon, a transitional water body located in the northeastern part of Italy, is discussed as a case study, by presenting the procedures adopted to measure the extent of the pollution, the impacts on organisms and the restoration activities