Novel Cell- and Tissue-Based Assays for Detecting Misfolded and Aggregated Protein Accumulation Within Aggresomes and Inclusion Bodies

Aggresomes and related inclusion bodies appear to serve as storage depots for misfolded and aggregated proteins within cells, which can potentially be degraded by the autophagy pathway. A homogenous fluorescence-based assay was devised to detect aggregated proteins inside aggresomes and inclusion bodies within an authentic cellular context. The assay employs a novel red fluorescent molecular rotor dye, which is essentially nonfluorescent until it binds to structural features associated with the aggregated protein cargo. Aggresomes and related structures were generated within cultured cells using various potent, cell permeable, proteasome inhibitors: MG-132, lactacystin, epoxomicin and bortezomib, and then selectively detected with the fluorescent probe. Employing the probe in combination with various fluorescein-labeled primary antibodies facilitated co-localization of key components of the autophagy system (ubiquitin, p62, and LC3) with aggregated protein cargo by fluorescence microscopy. Furthermore, cytoplasmic aggregates were highlighted in SK-N-SH human neuroblastoma cells incubated with exogenously supplied amyloid beta peptide 1–42. SMER28, a small molecule modulator of autophagy acting via an mTOR-independent mechanism, prevented the accumulation of amyloid beta peptide within these cells. The described assay allows assessment of the effects of protein aggregation directly in cells, without resorting to the use of non-physiological protein mutations or genetically engineered cell lines. With minor modification, the assay was also adapted to the analysis of frozen or formalin-fixed, paraffin-embedded tissue sections, with demonstration of co-localization of aggregated cargo with β-amyloid and tau proteins in brain tissue sections from Alzheimer’s disease patients

Cellular Communication through Light

Information transfer is a fundamental of life. A few studies have reported that cells use photons (from an endogenous source) as information carriers. This study finds that cells can have an influence on other cells even when separated with a glass barrier, thereby disabling molecule diffusion through the cell-containing medium. As there is still very little known about the potential of photons for intercellular communication this study is designed to test for non-molecule-based triggering of two fundamental properties of life: cell division and energy uptake. The study was performed with a cellular organism, the ciliate Paramecium caudatum. Mutual exposure of cell populations occurred under conditions of darkness and separation with cuvettes (vials) allowing photon but not molecule transfer. The cell populations were separated either with glass allowing photon transmission from 340 nm to longer waves, or quartz being transmittable from 150 nm, i.e. from UV-light to longer waves. Even through glass, the cells affected cell division and energy uptake in neighboring cell populations. Depending on the cuvette material and the number of cells involved, these effects were positive or negative. Also, while paired populations with lower growth rates grew uncorrelated, growth of the better growing populations was correlated. As there were significant differences when separating the populations with glass or quartz, it is suggested that the cell populations use two (or more) frequencies for cellular information transfer, which influences at least energy uptake, cell division rate and growth correlation. Altogether the study strongly supports a cellular communication system, which is different from a molecule-receptor-based system and hints that photon-triggering is a fine tuning principle in cell chemistry

Impact of Clinical Characteristics of Individual Metabolic Syndrome on the Severity of Insulin Resistance in Chinese Adults

The impact the metabolic syndrome (MetS) components on the severity of insulin resistance (IR) has not been reported. We enrolled 564 subjects with MetS and they were divided into quartiles according to the level of each component; and an insulin suppression test was performed to measure IR. In males, steady state plasma glucose (SSPG) levels in the highest quartiles, corresponding to body mass index (BMI) and fasting plasma glucose (FPG), were higher than the other three quartiles and the highest quartiles, corresponding to the diastolic blood pressure and triglycerides, were higher than in the lowest two quartiles. In females, SSPG levels in the highest quartiles, corresponding to the BMI and triglycerides, were higher than in all other quartiles. No significant differences existed between genders, other than the mean SSPG levels in males were greater in the highest quartile corresponding to BMI than that in the highest quartile corresponding to HDL-cholesterol levels. The factor analysis identified two underlying factors (IR and blood pressure factors) among the MetS variables. The clustering of the SSPG, BMI, triglyceride and HDL-cholesterol was noted. Our data suggest that adiposity, higher FPG and triglyceride levels have stronger correlation with IR and subjects with the highest BMI have the highest IR

Expression of genes for bone morphogenetic proteins BMP-2, BMP-4 and BMP-6 in various parts of the human skeleton

BACKGROUND: Differences in duration of bone healing in various parts of the human skeleton are common experience for orthopaedic surgeons. The reason for these differences is not obvious and not clear.METHODS: In this paper we decided to measure by the use of real-time RT-PCR technique the level of expression of genes for some isoforms of bone morphogenetic proteins (BMPs), whose role is proven in bone formation, bone induction and bone turnover. Seven bone samples recovered from various parts of skeletons from six cadavers of young healthy men who died in traffic accidents were collected. Activity of genes for BMP-2, -4 and -6 was measured by the use of fluorescent SYBR Green I.RESULTS: It was found that expression of m-RNA for BMP-2 and BMP-4 is higher in trabecular bone in epiphyses of long bones, cranial flat bones and corpus mandibulae then in the compact bone of diaphyses of long bones. In all samples examined the expression of m-RNA for BMP-4 was higher than for BMP-2.CONCLUSION: It was shown that m-RNA for BMP-6 is not expressed in the collected samples at all. It is postulated that differences in the level of activation of genes for BMPs is one of the important factors which determine the differences in duration of bone healing of various parts of the human skeleton.Author has checked copyrightDG 16/11/1

Achieving global biodiversity goals by 2050 requires urgent and integrated actions

Governments are negotiating actions intended to halt biodiversity loss and put it on a path to recovery by 2050. Here, we show that bending the curve for biodiversity is possible, but only if actions are implemented urgently and in an integrated manner. Connecting these actions to biodiversity outcomes and tracking progress remain a challenge

Statistical analysis of coverage error in simple global temperature estimators

Background Global mean surface temperature is widely used in the climate literature as a measure of the impact of human activity on the climate system. While the concept of a spatial average is simple, the estimation of that average from spatially incomplete data is not. Correlation between nearby map grid cells means that missing data cannot simply be ignored. Estimators that (often implicitly) assume uncorrelated observations can be biased when naively applied to the observed data, and in particular, the commonly used area weighted average is a biased estimator under these circumstances. Some surface temperature products use different forms of infilling or imputation to estimate temperatures for regions distant from the nearest observation, however the impacts of such methods on estimation of the global mean are not necessarily obvious or themselves unbiased. This issue was addressed in the 1970s by Ruvim Kagan, however his work has not been widely adopted, possibly due to its complexity and dependence on subjective choices in estimating the dependence between geographically proximate observations. Objectives The aim of this work is to present a simple estimator for global mean surface temperature from spatially incomplete data which retains many of the benefits of the work of Kagan, while being fully specified by two equations and a single parameter. The main purpose of the simplified estimator is to better explain to users of temperature data the problems associated with estimating an unbiased global mean from spatially incomplete data, however the estimator may also be useful for problems with specific requirements for reproducibility and performance. Methods The new estimator is based on generalized least squares, and uses the correlation matrix of the observations to weight each observation in accordance with the independent information it contributes. It can be implemented in fewer than 20 lines of computer code. The performance of the estimator is evaluated for different levels of observational coverage using reanalysis data with artificial noise. Results For recent decades the generalized least squares estimator mitigates most of the error associated with the use of a naive area weighted average. The improvement arises from the fact that coverage bias in the historical temperature record does not arise from an absolute shortage of observations (at least for recent decades), but rather from spatial heterogeneity in the distribution of observations, with some regions being relatively undersampled and others oversampled. The estimator addresses this problem by reducing the weight of the oversampled regions, in contrast to some existing global temperature datasets which extrapolate temperatures into the unobserved regions. The results are almost identical to the use of kriging (Gaussian process interpolation) to impute missing data to global coverage, followed by an area weighted average of the resulting field. However, the new formulation allows direct diagnosis of the contribution of individual observations and sources of error. Conclusions More sophisticated solutions to the problem of missing data in global temperature estimation already exist. However the simple estimator presented here, and the error analysis that it enables, demonstrate why such solutions are necessary. The 2013 Fifth Assessment Report of the Intergovernmental Panel on Climate Change discussed a slowdown in warming for the period 1998-2012, quoting the trend in the HadCRUT4 historical temperature dataset from the United Kingdom Meteorological Office in collaboration with the Climatic Research Unit of the University of East Anglia, along with other records. Use of the new estimator for global mean surface temperature would have reduced the apparent slowdown in warming of the early 21st century by one third in the spatially incomplete HadCRUT4 product

Transcriptional Landscape of the Prenatal Human Brain

Summary The anatomical and functional architecture of the human brain is largely determined by prenatal transcriptional processes. We describe an anatomically comprehensive atlas of mid-gestational human brain, including de novo reference atlases, in situ hybridization, ultra-high resolution magnetic resonance imaging (MRI) and microarray analysis on highly discrete laser microdissected brain regions. In developing cerebral cortex, transcriptional differences are found between different proliferative and postmitotic layers, wherein laminar signatures reflect cellular composition and developmental processes. Cytoarchitectural differences between human and mouse have molecular correlates, including species differences in gene expression in subplate, although surprisingly we find minimal differences between the inner and human-expanded outer subventricular zones. Both germinal and postmitotic cortical layers exhibit fronto-temporal gradients, with particular enrichment in frontal lobe. Finally, many neurodevelopmental disorder and human evolution-related genes show patterned expression, potentially underlying unique features of human cortical formation. These data provide a rich, freely-accessible resource for understanding human brain development

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

Recent Tropical Expansion: Natural Variability or Forced Response?

Previous studies have documented a poleward shift in the subsiding branches of Earth’s Hadley circulation since 1979 but have disagreed on the causes of these observed changes and the ability of global climate models to capture them. This synthesis paper reexamines a number of contradictory claims in the past literature and finds that the tropical expansion indicated by modern reanalyses is within the bounds of models’ historical simulations for the period 1979–2005. Earlier conclusions that models were underestimating the observed trends relied on defining the Hadley circulation using the mass streamfunction from older reanalyses. The recent observed tropical expansion has similar magnitudes in the annual mean in the Northern Hemisphere (NH) and Southern Hemisphere (SH), but models suggest that the factors driving the expansion differ between the hemispheres. In the SH, increasing greenhouse gases (GHGs) and stratospheric ozone depletion contributed to tropical expansion over the late twentieth century, and if GHGs continue increasing, the SH tropical edge is projected to shift further poleward over the twenty-first century, even as stratospheric ozone concentrations recover. In the NH, the contribution of GHGs to tropical expansion is much smaller and will remain difficult to detect in a background of large natural variability, even by the end of the twenty-first century. To explain similar recent tropical expansion rates in the two hemispheres, natural variability must be taken into account. Recent coupled atmosphere–ocean variability, including the Pacific decadal oscillation, has contributed to tropical expansion. However, in models forced with observed sea surface temperatures, tropical expansion rates still vary widely because of internal atmospheric variability