MicroRNAs (miRNA) are small non-coding RNAs that regulate translation of mRNA and protein. Loss or enhanced expression of miRNAs is associated with several diseases, including cancer. However, the ...identification of circulating miRNA in healthy donors is not well characterized. Microvesicles, also known as exosomes or microparticles, circulate in the peripheral blood and can stimulate cellular signaling. In this study, we hypothesized that under normal healthy conditions, microvesicles contain miRNAs, contributing to biological homeostasis.
Microvesicles were isolated from the plasma of normal healthy individuals. RNA was isolated from both the microvesicles and matched mononuclear cells and profiled for 420 known mature miRNAs by real-time PCR. Hierarchical clustering of the data sets indicated significant differences in miRNA expression between peripheral blood mononuclear cells (PBMC) and plasma microvesicles. We observed 71 miRNAs co-expressed between microvesicles and PBMC. Notably, we found 33 and 4 significantly differentially expressed miRNAs in the plasma microvesicles and mononuclear cells, respectively. Prediction of the gene targets and associated biological pathways regulated by the detected miRNAs was performed. The majority of the miRNAs expressed in the microvesicles from the blood were predicted to regulate cellular differentiation of blood cells and metabolic pathways. Interestingly, a select few miRNAs were also predicted to be important modulators of immune function.
This study is the first to identify and define miRNA expression in circulating plasma microvesicles of normal subjects. The data generated from this study provides a basis for future studies to determine the predictive role of peripheral blood miRNA signatures in human disease and will enable the definition of the biological processes regulated by these miRNA.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
MicroRNAs (miRNAs) have emerged as important regulators in the post‐transcriptional control of gene expression. The discovery of their presence not only in tissues but also in extratissular fluids, ...including blood, urine and cerebro‐spinal fluid, together with their changes in expression in various pathological conditions, has implicated these extracellular miRNAs as informative biomarkers of disease. However, exploiting miRNAs in this capacity requires methodological rigour. Here, we report several key procedural aspects of miRNA isolation from plasma and serum, as exemplified by research in cardiovascular and pulmonary diseases. We also highlight the advantages and disadvantages of various profiling methods to determine the expression levels of plasma‐ and serum‐derived miRNAs. Attention to such methodological details is critical, as circulating miRNAs become diagnostic tools for various human diseases.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
The homogeneous coordinate ring of the Grassmannian Gr
k
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n
has a cluster structure defined in terms of planar diagrams known as Postnikov diagrams. The cluster corresponding to such a diagram ...consists entirely of Plücker coordinates. We introduce a twist map on Gr
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, related to the Berenstein–Fomin–Zelevinsky-twist, and give an explicit Laurent expansion for the twist of an arbitrary Plücker coordinate in terms of the cluster variables associated with a fixed Postnikov diagram. The expansion arises as a (scaled) dimer partition function of a weighted version of the bipartite graph dual to the Postnikov diagram, modified by a boundary condition determined by the Plücker coordinate. We also relate the twist map to a maximal green sequence.
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DOBA, EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NLZOH, NUK, ODKLJ, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, SIK, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
The role of inflammation in idiopathic pulmonary fibrosis (IPF) is controversial. If inflammation were critical to the disease process, lung pathology would demonstrate an influx of inflammatory ...cells, and that the disease would respond to immunosuppression. Neither is true. The classic pathology does not display substantial inflammation, and no modulation of the immune system is effective as treatment. Recent data suggest that the pathophysiology of the disease is more a product of fibroblast dysfunction than of dysregulated inflammation. The role of inflammation in disease pathogenesis comes from pathology from atypical patients, biologic samples procured during exacerbations of the disease, and careful examination of biologic specimens from patients with stable disease. We suggest that inflammation is indeed a critical factor in IPF and propose five potential nontraditional mechanisms for the role of inflammation in the pathogenesis of IPF: the direct inflammatory hypothesis, the matrix hypothesis, the growth factor-receptor hypothesis, the plasticity hypothesis, and the vascular hypothesis. To address these, we review the literature exploring the differences in pathology, prognosis, and clinical course, as well as the role of cytokines, growth factors, and other mediators of inflammation, and last, the role of matrix and vascular supply in patients with IPF.
Altered inflammatory cytokine profiles are often observed in individuals suffering from major depression. Recent clinical work reports on elevated IL-6 and decreased IL-10 in depression. Elevated ...IL-6 has served as a consistent biomarker of depression and IL-10 is proposed to influence depressive behavior through its ability to counterbalance pro-inflammatory cytokine expression. Clinical and animal studies suggest a role for IL-10 in modifying depressive behavior. Murine restraint stress (RST) is regularly employed in the study of behavioral and biological symptoms associated with depressive disorders. While responses to acute RST exposure have been widely characterized, few studies have examined the ongoing and longitudinal effects of extended RST and fewer still have examined the lasting impact during the post-stress period. Consistent with clinical data, we report that a protocol of prolonged murine RST produced altered cytokine profiles similar to those observed in major depressive disorder. Parallel to these changes in circulating cytokines, IL-10 mRNA expression was diminished in the cortex and hippocampus throughout the stress period and following cessation of RST. Moreover, chronic RST promoted depressive-like behavior throughout the 28-day stress period and these depressive-like complications were maintained weeks after cessation of RST. Because of the correlation between IL-10 suppression and depressive behavior and because many successful antidepressant therapies yield increases in IL-10, we examined the effects of IL-10 treatment on RST-induced behavioral changes. Behavioral deficits induced by RST were reversed by exogenous administration of recombinant IL-10. This work provides one of the first reports describing the biological and behavioral impact following prolonged RST and, taken together, this study provides details on the correlation between responses to chronic RST and those seen in depressive disorders.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The transcription factors E2F and Myc participate in the control of cell proliferation and apoptosis, and can act as oncogenes or tumor suppressors depending on their levels of expression. Positive ...feedback loops in the regulation of these factors are predicted--and recently shown experimentally--to lead to bistability, which is a phenomenon characterized by the existence of low and high protein levels ("off" and "on" levels, respectively), with sharp transitions between levels being inducible by, for example, changes in growth factor concentrations. E2F and Myc are inhibited at the posttranscriptional step by members of a cluster of microRNAs (miRs) called miR-17-92. In return, E2F and Myc induce the transcription of miR-17-92, thus forming a negative feedback loop in the interaction network. The consequences of the coupling between the E2F/Myc positive feedback loops and the E2F/Myc/miR-17-92 negative feedback loop are analyzed using a mathematical model. The model predicts that miR-17-92 plays a critical role in regulating the position of the off-on switch in E2F/Myc protein levels, and in determining the on levels of these proteins. The model also predicts large-amplitude protein oscillations that coexist with the off steady state levels. Using the concept and model prediction of a "cancer zone," the oncogenic and tumor suppressor properties of miR-17-92 is demonstrated to parallel the same properties of E2F and Myc.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Abstract
Background
Quantitative PCR (qPCR) aims to measure the DNA or RNA concentration in diagnostic and biological samples based on the quantification cycle (Cq) value observed in the ...amplification curves. Results of qPCR experiments are regularly calculated as if all assays are 100% efficient or reported as just Cq, ΔCq, or ΔΔCq values.
Contents
When the reaction shows specific amplification, it should be deemed to be positive, regardless of the observed Cq. Because the Cq is highly dependent on amplification efficiency that can vary among targets and samples, accurate calculation of the target quantity and relative gene expression requires that the actual amplification efficiency be taken into account in the analysis and reports. PCR efficiency is frequently derived from standard curves, but this approach is affected by dilution errors and hampered by properties of the standard and the diluent. These factors affect accurate quantification of clinical and biological samples used in diagnostic applications and collected in challenging conditions. PCR efficiencies determined from individual amplification curves avoid these confounders. To obtain unbiased efficiency-corrected results, we recommend absolute quantification with a single undiluted calibrator with a known target concentration and efficiency values derived from the amplification curves of the calibrator and the unknown samples.
Summary
For meaningful diagnostics or biological interpretation, the reported results of qPCR experiments should be efficiency corrected. To avoid ambiguity, the Minimal Information for Publications on Quantitative Real-Time PCR Experiments (MIQE) guidelines checklist should be extended to require the methods that were used (1) to determine the PCR efficiency and (2) to calculate the reported target quantity and relative gene expression value.
Estimates of near‐surface wind speed and direction are key meteorological components for predicting many surface hydrometeorological processes that influence critical aspects of hydrological and ...biological systems. However, observations of near‐surface wind are typically spatially sparse. The use of these sparse wind fields to force distributed models, such as hydrological models, is greatly complicated in complex terrain, such as mountain headwaters basins. In these regions, wind flows are heavily impacted by overlapping influences of terrain at different scales. This can have a great impact on calculations of evapotranspiration, snowmelt, and blowing snow transport and sublimation. The use of high‐resolution atmospheric models allows for numerical weather prediction (NWP) model outputs to be dynamically downscaled. However, the computation burden for large spatial extents and long periods of time often precludes their use. Here, a wind‐library approach is presented to aid in downscaling NWP outputs and terrain‐correcting spatially interpolated observations. This approach preserves important spatial characteristics of the flow field at a fraction of the computational costs of even the simplest high‐resolution atmospheric models. This approach improves on previous implementations by: scaling to large spatial extents O(1M km2); approximating lee‐side effects; and fully automating the creation of the wind library. Overall, this approach was shown to have a third quartile RMSE of 1.8 m·s−1 $\mathrm{m}\cdot {\mathrm{s}}^{-1}$ and a third quartile RMSE of 58.2° versus a standalone diagnostic windflow model. The wind velocity estimates versus observations were better than existing empirical terrain‐based estimates and computational savings were approximately 100‐fold versus the diagnostic model.
Key Points
New software for wind library generation
Lower error in speed and direction that other empirical models
Applicable to large extents over 1M km2
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Idiopathic pulmonary fibrosis (IPF) is a disease of progressive lung fibrosis with a high mortality rate. In organ repair and remodeling, epigenetic events are important. MicroRNAs (miRNAs) regulate ...gene expression post-transcriptionally and can target epigenetic molecules important in DNA methylation. The miR-17~92 miRNA cluster is critical for lung development and lung epithelial cell homeostasis and is predicted to target fibrotic genes and DNA methyltransferase (DNMT)-1 expression.
We investigated the miR-17~92 cluster expression and its role in regulating DNA methylation events in IPF lung tissue.
Expression and DNA methylation patterns of miR-17~92 were determined in human IPF lung tissue and fibroblasts and fibrotic mouse lung tissue. The relationship between the miR-17~92 cluster and DNMT-1 expression was examined in vitro. Using a murine model of pulmonary fibrosis, we examined the therapeutic potential of the demethylating agent, 5'-aza-2'-deoxycytidine.
Compared with control samples, miR-17~92 expression was reduced in lung biopsies and lung fibroblasts from patients with IPF, whereas DNMT-1 expression and methylation of the miR-17~92 promoter was increased. Several miRNAs from the miR-17~92 cluster targeted DNMT-1 expression resulting in a negative feedback loop. Similarly, miR-17~92 expression was reduced in the lungs of bleomycin-treated mice. Treatment with 5'-aza-2'-deoxycytidine in a murine bleomycin-induced pulmonary fibrosis model reduced fibrotic gene and DNMT-1 expression, enhanced miR-17~92 cluster expression, and attenuated pulmonary fibrosis.
This study provides insight into the pathobiology of IPF and identifies a novel epigenetic feedback loop between miR-17~92 and DNMT-1 in lung fibrosis.
The interaction of mountain terrain with meteorological processes causes substantial temporal and spatial variability in snow accumulation and ablation. Processes impacted by complex terrain include ...large-scale orographic enhancement of snowfall, small-scale processes such as gravitational and wind-induced transport of snow, and variability in the radiative balance such as through terrain shadowing. In this study, a multi-scale modelling approach is proposed to simulate the temporal and spatial evolution of high-mountain snowpacks. The multi-scale approach combines atmospheric data from a numerical weather prediction system at the kilometre scale with process-based downscaling techniques to drive the Canadian Hydrological Model (CHM) at spatial resolutions allowing for explicit snow redistribution modelling. CHM permits a variable spatial resolution by using the efficient terrain representation by unstructured triangular meshes. The model simulates processes such as radiation shadowing and irradiance to slopes, blowing-snow transport (saltation and suspension) and sublimation, avalanching, forest canopy interception and sublimation, and snowpack melt. Short-term, kilometre-scale atmospheric forecasts from Environment and Climate Change Canada's Global Environmental Multiscale Model through its High Resolution Deterministic Prediction System (HRDPS) drive CHM and are downscaled to the unstructured mesh scale. In particular, a new wind-downscaling strategy uses pre-computed wind fields from a mass-conserving wind model at 50 m resolution to perturb the mesoscale HRDPS wind and to account for the influence of topographic features on wind direction and speed. HRDPS-CHM was applied to simulate snow conditions down to 50 m resolution during winter 2017/2018 in a domain around the Kananaskis Valley (∼1000 km2) in the Canadian Rockies. Simulations were evaluated using high-resolution airborne light detection and ranging (lidar) snow depth data and snow persistence indexes derived from remotely sensed imagery. Results included model falsifications and showed that both wind-induced and gravitational snow redistribution need to be simulated to capture the snowpack variability and the evolution of snow depth and persistence with elevation across the region. Accumulation of windblown snow on leeward slopes and associated snow cover persistence were underestimated in a CHM simulation driven by wind fields that did not capture lee-side flow recirculation and associated wind speed decreases. A terrain-based metric helped to identify these lee-side areas and improved the wind field and the associated snow redistribution. An overestimation of snow redistribution from windward to leeward slopes and subsequent avalanching was still found. The results of this study highlight the need for further improvements of snowdrift-permitting models for large-scale applications, in particular the representation of subgrid topographic effects on snow transport.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
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