Cross-correlation analysis to quantify relative spatial distributions of fat and protein in super-resolution microscopy images of dairy gels

The advent of super-resolution microscopy allows microstructures of foods to be explored in new depths, which when coupled with quantitative image analysis can provide a powerful analytical tool. Herein, a methodology is presented and applied to use a 2D spatial cross-correlation analysis to investigate the relative spatial arrangement of protein and fat in acid induced whole milk gels where the milk is either non-homogenised or has been homogenised at either 10 or 25 MPa. Two-channel images were taken using super-resolution Stimulated Emission Depletion (STED) microscopy and confocal microscopy. A term has been derived to extract the typical distance from the fat droplet surface and to the local maximum protein distribution. The fat droplet size is determined through 2D spatial autocorrelation analysis. Methods of analysis are applied to global images and to region specific analysis focussing on individual fat droplets. Cross-correlation analysis has been empirically validated using generated images with precise spatial features corresponding to the features of interest in true microscopy images, over appropriate length scales. The protein microstructure, fat droplet size and distances between the fat droplets and protein network are characterised. There are significantly different distances between the fat droplets and protein network in the homogenised samples compared to the non-homogenised sample. The extracted separation distances are below the diffraction limit of light, highlighting the utility of super-resolution imaging

Dynamic moisture loss explored through quantitative super-resolution microscopy, spatial micro-viscosity and macroscopic analyses in acid milk gels

Molecular interactions and dynamic changes at a range of length scales affect the structuring of food materials, as such it is essential to explore structure at a range of different length scales. Herein, four acid milk gel samples are produced from either fresh or reconstituted skim milk that either had no heat treatment or had undergone heat treatment at 85 °C for 10 min. Milk acid gels demonstrate complex structure on a range of length scales of interest in colloidal materials and exhibit different macroscopic and water binding properties. A method is presented to measure the dynamic moisture loss in these samples, without applying external force. Super-resolution microscopy images are quantitatively analysed to describe the gel microstructure with precise features. Fluorescent Lifetime Imaging Microscopy is used to spatially resolve differences in molecular confinement across the sample's microstructure, which is quantified for each sample. Moisture loss and microstructural analyses are correlated to bulk and macroscopic properties determined through rheological and texture analysis, pH and conductivity measurements. More severe thermal and processing treatments leads to a reduction in moisture loss over time. Differences in moisture loss and mechanical properties relate to different thermal processing histories, but are not fully explained by levels of denatured whey proteins, and appear related to changes in mineral balance. The methods presented provide a comprehensive and complementary overview of material properties across relevant length scales and relevant sample conditions

Simulation Modifies Prehension: Evidence for a Conjoined Representation of the Graspable Features of an Object and the Action of Grasping It

Movement formulas, engrams, kinesthetic images and internal models of the body in action are notions derived mostly from clinical observations of brain-damaged subjects. They also suggest that the prehensile geometry of an object is integrated in the neural circuits and includes the object's graspable characteristics as well as its semantic properties. In order to determine whether there is a conjoined representation of the graspable characteristics of an object in relation to the actual grasping, it is necessary to separate the graspable (low-level) from the semantic (high-level) properties of the object. Right-handed subjects were asked to grasp and lift a smooth 300-g cylinder with one hand, before and after judging the level of difficulty of a “grasping for pouring” action, involving a smaller cylinder and using the opposite hand. The results showed that simulated grasps with the right hand exert a direct influence on actual motor acts with the left hand. These observations add to the evidence that there is a conjoined representation of the graspable characteristics of the object and the biomechanical constraints of the arm

Perfect Hash Families: The Generalization to Higher Indices

Perfect hash families are often represented as combinatorial arrays encoding partitions of kitems into v classes, so that every t or fewer of the items are completely separated by at least a specified number of chosen partitions. This specified number is the index of the hash family. The case when each t-set must be separated at least once has been extensively researched; they arise in diverse applications, both directly and as fundamental ingredients in a column replacement strategy for a variety of combinatorial arrays. In this paper, construction techniques and algorithmic methods for constructing perfect hash families are surveyed, in order to explore extensions to the situation when each t-set must be separated by more than one partition.https://digitalcommons.usmalibrary.org/books/1029/thumbnail.jp

RNA Polymerase II Pausing Downstream of Core Histone Genes Is Different from Genes Producing Polyadenylated Transcripts

Recent genome-wide chromatin immunoprecipitation coupled high throughput sequencing (ChIP-seq) analyses performed in various eukaryotic organisms, analysed RNA Polymerase II (Pol II) pausing around the transcription start sites of genes. In this study we have further investigated genome-wide binding of Pol II downstream of the 3′ end of the annotated genes (EAGs) by ChIP-seq in human cells. At almost all expressed genes we observed Pol II occupancy downstream of the EAGs suggesting that Pol II pausing 3′ from the transcription units is a rather common phenomenon. Downstream of EAGs Pol II transcripts can also be detected by global run-on and sequencing, suggesting the presence of functionally active Pol II. Based on Pol II occupancy downstream of EAGs we could distinguish distinct clusters of Pol II pause patterns. On core histone genes, coding for non-polyadenylated transcripts, Pol II occupancy is quickly dropping after the EAG. In contrast, on genes, whose transcripts undergo polyA tail addition [poly(A)+], Pol II occupancy downstream of the EAGs can be detected up to 4–6 kb. Inhibition of polyadenylation significantly increased Pol II occupancy downstream of EAGs at poly(A)+ genes, but not at the EAGs of core histone genes. The differential genome-wide Pol II occupancy profiles 3′ of the EAGs have also been confirmed in mouse embryonic stem (mES) cells, indicating that Pol II pauses genome-wide downstream of the EAGs in mammalian cells. Moreover, in mES cells the sharp drop of Pol II signal at the EAG of core histone genes seems to be independent of the phosphorylation status of the C-terminal domain of the large subunit of Pol II. Thus, our study uncovers a potential link between different mRNA 3′ end processing mechanisms and consequent Pol II transcription termination processes

Plant 45S rDNA Clusters Are Fragile Sites and Their Instability Is Associated with Epigenetic Alterations

Our previous study demonstrated that 45S ribosomal DNA (45S rDNA) clusters were chromosome fragile sites expressed spontaneously in Lolium. In this study, fragile phenotypes of 45S rDNA were observed under aphidicolin (APH) incubation in several plant species. Further actinomycin D (ActD) treatment showed that transcriptional stress might interfere with chromatin packaging, resulting in 45S rDNA fragile expression. These data identified 45S rDNA sites as replication-dependent as well as transcription-dependent fragile sites in plants. In the presence of ActD, a dramatic switch to an open chromatin conformation and accumulated incomplete 5′ end of the external transcribed spacer (5′ETS) transcripts were observed, accompanied by decreased DNA methylation, decreased levels of histone H3, and increased histone acetylation and levels of H3K4me2, suggesting that these epigenetic alterations are associated with failure of 45S rDNA condensation. Furthermore, the finding that γ-H2AX was accumulated at 45S rDNA sites following ActD treatment suggested that the DNA damage signaling pathway was associated with the appearance of 45S rDNA fragile phenotypes. Our data provide a link between 45S rDNA transcription and chromatin-packaging defects and open the door for further identifying the molecular mechanism involved

Cardiovascular magnetic resonance in pericardial diseases

The pericardium and pericardial diseases in particular have received, in contrast to other topics in the field of cardiology, relatively limited interest. Today, despite improved knowledge of pathophysiology of pericardial diseases and the availability of a wide spectrum of diagnostic tools, the diagnostic challenge remains. Not only the clinical presentation may be atypical, mimicking other cardiac, pulmonary or pleural diseases; in developed countries a shift for instance in the epidemiology of constrictive pericarditis has been noted. Accurate decision making is crucial taking into account the significant morbidity and mortality caused by complicated pericardial diseases, and the potential benefit of therapeutic interventions. Imaging herein has an important role, and cardiovascular magnetic resonance (CMR) is definitely one of the most versatile modalities to study the pericardium. It fuses excellent anatomic detail and tissue characterization with accurate evaluation of cardiac function and assessment of the haemodynamic consequences of pericardial constraint on cardiac filling. This review focuses on the current state of knowledge how CMR can be used to study the most common pericardial diseases

Host-parasite co-metabolic activation of antitrypanosomal aminomethyl-benzoxaboroles

<div><p>Recent development of benzoxaborole-based chemistry gave rise to a collection of compounds with great potential in targeting diverse infectious diseases, including human African Trypanosomiasis (HAT), a devastating neglected tropical disease. However, further medicinal development is largely restricted by a lack of insight into mechanism of action (MoA) in pathogenic kinetoplastids. We adopted a multidisciplinary approach, combining a high-throughput forward genetic screen with functional group focused chemical biological, structural biology and biochemical analyses, to tackle the complex MoAs of benzoxaboroles in <i>Trypanosoma brucei</i>. We describe an oxidative enzymatic pathway composed of host semicarbazide-sensitive amine oxidase and a trypanosomal aldehyde dehydrogenase TbALDH3. Two sequential reactions through this pathway serve as the key underlying mechanism for activating a series of 4-aminomethylphenoxy-benzoxaboroles as potent trypanocides; the methylamine parental compounds as pro-drugs are transformed first into intermediate aldehyde metabolites, and further into the carboxylate metabolites as effective forms. Moreover, comparative biochemical and crystallographic analyses elucidated the catalytic specificity of TbALDH3 towards the benzaldehyde benzoxaborole metabolites as xenogeneic substrates. Overall, this work proposes a novel drug activation mechanism dependent on both host and parasite metabolism of primary amine containing molecules, which contributes a new perspective to our understanding of the benzoxaborole MoA, and could be further exploited to improve the therapeutic index of antimicrobial compounds.</p></div

Antiinflammatory Therapy with Canakinumab for Atherosclerotic Disease

Background: Experimental and clinical data suggest that reducing inflammation without affecting lipid levels may reduce the risk of cardiovascular disease. Yet, the inflammatory hypothesis of atherothrombosis has remained unproved. Methods: We conducted a randomized, double-blind trial of canakinumab, a therapeutic monoclonal antibody targeting interleukin-1β, involving 10,061 patients with previous myocardial infarction and a high-sensitivity C-reactive protein level of 2 mg or more per liter. The trial compared three doses of canakinumab (50 mg, 150 mg, and 300 mg, administered subcutaneously every 3 months) with placebo. The primary efficacy end point was nonfatal myocardial infarction, nonfatal stroke, or cardiovascular death. RESULTS: At 48 months, the median reduction from baseline in the high-sensitivity C-reactive protein level was 26 percentage points greater in the group that received the 50-mg dose of canakinumab, 37 percentage points greater in the 150-mg group, and 41 percentage points greater in the 300-mg group than in the placebo group. Canakinumab did not reduce lipid levels from baseline. At a median follow-up of 3.7 years, the incidence rate for the primary end point was 4.50 events per 100 person-years in the placebo group, 4.11 events per 100 person-years in the 50-mg group, 3.86 events per 100 person-years in the 150-mg group, and 3.90 events per 100 person-years in the 300-mg group. The hazard ratios as compared with placebo were as follows: in the 50-mg group, 0.93 (95% confidence interval [CI], 0.80 to 1.07; P = 0.30); in the 150-mg group, 0.85 (95% CI, 0.74 to 0.98; P = 0.021); and in the 300-mg group, 0.86 (95% CI, 0.75 to 0.99; P = 0.031). The 150-mg dose, but not the other doses, met the prespecified multiplicity-adjusted threshold for statistical significance for the primary end point and the secondary end point that additionally included hospitalization for unstable angina that led to urgent revascularization (hazard ratio vs. placebo, 0.83; 95% CI, 0.73 to 0.95; P = 0.005). Canakinumab was associated with a higher incidence of fatal infection than was placebo. There was no significant difference in all-cause mortality (hazard ratio for all canakinumab doses vs. placebo, 0.94; 95% CI, 0.83 to 1.06; P = 0.31). Conclusions: Antiinflammatory therapy targeting the interleukin-1β innate immunity pathway with canakinumab at a dose of 150 mg every 3 months led to a significantly lower rate of recurrent cardiovascular events than placebo, independent of lipid-level lowering. (Funded by Novartis; CANTOS ClinicalTrials.gov number, NCT01327846.

Multi-messenger observations of a binary neutron star merger

On 2017 August 17 a binary neutron star coalescence candidate (later designated GW170817) with merger time 12:41:04 UTC was observed through gravitational waves by the Advanced LIGO and Advanced Virgo detectors. The Fermi Gamma-ray Burst Monitor independently detected a gamma-ray burst (GRB 170817A) with a time delay of ~1.7 s with respect to the merger time. From the gravitational-wave signal, the source was initially localized to a sky region of 31 deg2 at a luminosity distance of 40+8-8 Mpc and with component masses consistent with neutron stars. The component masses were later measured to be in the range 0.86 to 2.26 Mo. An extensive observing campaign was launched across the electromagnetic spectrum leading to the discovery of a bright optical transient (SSS17a, now with the IAU identification of AT 2017gfo) in NGC 4993 (at ~40 Mpc) less than 11 hours after the merger by the One- Meter, Two Hemisphere (1M2H) team using the 1 m Swope Telescope. The optical transient was independently detected by multiple teams within an hour. Subsequent observations targeted the object and its environment. Early ultraviolet observations revealed a blue transient that faded within 48 hours. Optical and infrared observations showed a redward evolution over ~10 days. Following early non-detections, X-ray and radio emission were discovered at the transient’s position ~9 and ~16 days, respectively, after the merger. Both the X-ray and radio emission likely arise from a physical process that is distinct from the one that generates the UV/optical/near-infrared emission. No ultra-high-energy gamma-rays and no neutrino candidates consistent with the source were found in follow-up searches. These observations support the hypothesis that GW170817 was produced by the merger of two neutron stars in NGC4993 followed by a short gamma-ray burst (GRB 170817A) and a kilonova/macronova powered by the radioactive decay of r-process nuclei synthesized in the ejecta