3D time series analysis of cell shape using Laplacian approaches

Background: Fundamental cellular processes such as cell movement, division or food uptake critically depend on cells being able to change shape. Fast acquisition of three-dimensional image time series has now become possible, but we lack efficient tools for analysing shape deformations in order to understand the real three-dimensional nature of shape changes. Results: We present a framework for 3D+time cell shape analysis. The main contribution is three-fold: First, we develop a fast, automatic random walker method for cell segmentation. Second, a novel topology fixing method is proposed to fix segmented binary volumes without spherical topology. Third, we show that algorithms used for each individual step of the analysis pipeline (cell segmentation, topology fixing, spherical parameterization, and shape representation) are closely related to the Laplacian operator. The framework is applied to the shape analysis of neutrophil cells. Conclusions: The method we propose for cell segmentation is faster than the traditional random walker method or the level set method, and performs better on 3D time-series of neutrophil cells, which are comparatively noisy as stacks have to be acquired fast enough to account for cell motion. Our method for topology fixing outperforms the tools provided by SPHARM-MAT and SPHARM-PDM in terms of their successful fixing rates. The different tasks in the presented pipeline for 3D+time shape analysis of cells can be solved using Laplacian approaches, opening the possibility of eventually combining individual steps in order to speed up computations

Extracellular Matrix Aggregates from Differentiating Embryoid Bodies as a Scaffold to Support ESC Proliferation and Differentiation

Embryonic stem cells (ESCs) have emerged as potential cell sources for tissue engineering and regeneration owing to its virtually unlimited replicative capacity and the potential to differentiate into a variety of cell types. Current differentiation strategies primarily involve various growth factor/inducer/repressor concoctions with less emphasis on the substrate. Developing biomaterials to promote stem cell proliferation and differentiation could aid in the realization of this goal. Extracellular matrix (ECM) components are important physiological regulators, and can provide cues to direct ESC expansion and differentiation. ECM undergoes constant remodeling with surrounding cells to accommodate specific developmental event. In this study, using ESC derived aggregates called embryoid bodies (EB) as a model, we characterized the biological nature of ECM in EB after exposure to different treatments: spontaneously differentiated and retinoic acid treated (denoted as SPT and RA, respectively). Next, we extracted this treatment-specific ECM by detergent decellularization methods (Triton X-100, DOC and SDS are compared). The resulting EB ECM scaffolds were seeded with undifferentiated ESCs using a novel cell seeding strategy, and the behavior of ESCs was studied. Our results showed that the optimized protocol efficiently removes cells while retaining crucial ECM and biochemical components. Decellularized ECM from SPT EB gave rise to a more favorable microenvironment for promoting ESC attachment, proliferation, and early differentiation, compared to native EB and decellularized ECM from RA EB. These findings suggest that various treatment conditions allow the formulation of unique ESC-ECM derived scaffolds to enhance ESC bioactivities, including proliferation and differentiation for tissue regeneration applications. © 2013 Goh et al

BCAA catabolism in brown fat controls energy homeostasis through SLC25A44.

Branched-chain amino acid (BCAA; valine, leucine and isoleucine) supplementation is often beneficial to energy expenditure; however, increased circulating levels of BCAA are linked to obesity and diabetes. The mechanisms of this paradox remain unclear. Here we report that, on cold exposure, brown adipose tissue (BAT) actively utilizes BCAA in the mitochondria for thermogenesis and promotes systemic BCAA clearance in mice and humans. In turn, a BAT-specific defect in BCAA catabolism attenuates systemic BCAA clearance, BAT fuel oxidation and thermogenesis, leading to diet-induced obesity and glucose intolerance. Mechanistically, active BCAA catabolism in BAT is mediated by SLC25A44, which transports BCAAs into mitochondria. Our results suggest that BAT serves as a key metabolic filter that controls BCAA clearance via SLC25A44, thereby contributing to the improvement of metabolic health

Mesoporous monoliths of inverse bicontinuous cubic phases of block copolymer bilayers

Solution self-assembly of block copolymers into inverse bicontinuous cubic mesophases is a promising new approach for creating porous polymer films and monoliths with highly organized bicontinuous mesoporous networks. Here we report the direct self-assembly of block copolymers with branched hydrophilic blocks into large monoliths consisting of the inverse bicontinuous cubic structures of the block copolymer bilayer. We suggest a facile and scalable method of solution self-assembly by diffusion of water to the block copolymer solution, which results in the unperturbed formation of mesoporous monoliths with large-pore (>25nm diameter) networks weaved in crystalline lattices. The surface functional groups of the internal large-pore networks are freely accessible for large guest molecules such as protein complexes of which the molecular weight exceeded 100kDa. The internal double-diamond (Pn3m) networks of large pores within the mesoporous monoliths could be replicated to self-supporting three-dimensional skeletal structures of crystalline titania and mesoporous silica.open2

Quantitative utilization of prior biological knowledge in the Bayesian network modeling of gene expression data

<p>Abstract</p> <p>Background</p> <p>Bayesian Network (BN) is a powerful approach to reconstructing genetic regulatory networks from gene expression data. However, expression data by itself suffers from high noise and lack of power. Incorporating prior biological knowledge can improve the performance. As each type of prior knowledge on its own may be incomplete or limited by quality issues, integrating multiple sources of prior knowledge to utilize their consensus is desirable.</p> <p>Results</p> <p>We introduce a new method to incorporate the quantitative information from multiple sources of prior knowledge. It first uses the Naïve Bayesian classifier to assess the likelihood of functional linkage between gene pairs based on prior knowledge. In this study we included cocitation in PubMed and schematic similarity in Gene Ontology annotation. A candidate network edge reservoir is then created in which the copy number of each edge is proportional to the estimated likelihood of linkage between the two corresponding genes. In network simulation the Markov Chain Monte Carlo sampling algorithm is adopted, and samples from this reservoir at each iteration to generate new candidate networks. We evaluated the new algorithm using both simulated and real gene expression data including that from a yeast cell cycle and a mouse pancreas development/growth study. Incorporating prior knowledge led to a ~2 fold increase in the number of known transcription regulations recovered, without significant change in false positive rate. In contrast, without the prior knowledge BN modeling is not always better than a random selection, demonstrating the necessity in network modeling to supplement the gene expression data with additional information.</p> <p>Conclusion</p> <p>our new development provides a statistical means to utilize the quantitative information in prior biological knowledge in the BN modeling of gene expression data, which significantly improves the performance.</p

A Functional Gene Array for Detection of Bacterial Virulence Elements

Emerging known and unknown pathogens create profound threats to public health. Platforms for rapid detection and characterization of microbial agents are critically needed to prevent and respond to disease outbreaks. Available detection technologies cannot provide broad functional information about known or novel organisms. As a step toward developing such a system, we have produced and tested a series of high-density functional gene arrays to detect elements of virulence and antibiotic resistance mechanisms. Our first generation array targets genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for gene family detection and discrimination. When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family. In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms. This is the first report of a high density NimbleGen microarray system targeting microbial antibiotic resistance and virulence mechanisms. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples

Metagenomics of the Svalbard Reindeer Rumen Microbiome Reveals Abundance of Polysaccharide Utilization Loci

Lignocellulosic biomass remains a largely untapped source of renewable energy predominantly due to its recalcitrance and an incomplete understanding of how this is overcome in nature. We present here a compositional and comparative analysis of metagenomic data pertaining to a natural biomass-converting ecosystem adapted to austere arctic nutritional conditions, namely the rumen microbiome of Svalbard reindeer (Rangifer tarandus platyrhynchus). Community analysis showed that deeply-branched cellulolytic lineages affiliated to the Bacteroidetes and Firmicutes are dominant, whilst sequence binning methods facilitated the assemblage of metagenomic sequence for a dominant and novel Bacteroidales clade (SRM-1). Analysis of unassembled metagenomic sequence as well as metabolic reconstruction of SRM-1 revealed the presence of multiple polysaccharide utilization loci-like systems (PULs) as well as members of more than 20 glycoside hydrolase and other carbohydrate-active enzyme families targeting various polysaccharides including cellulose, xylan and pectin. Functional screening of cloned metagenome fragments revealed high cellulolytic activity and an abundance of PULs that are rich in endoglucanases (GH5) but devoid of other common enzymes thought to be involved in cellulose degradation. Combining these results with known and partly re-evaluated metagenomic data strongly indicates that much like the human distal gut, the digestive system of herbivores harbours high numbers of deeply branched and as-yet uncultured members of the Bacteroidetes that depend on PUL-like systems for plant biomass degradation

Anogenital distance in human male and female newborns: a descriptive, cross-sectional study

BACKGROUND: In animal studies of the effects of hormonally active agents, measurement of anogenital distance (AGD) is now routine, and serves as a bioassay of fetal androgen action. Although measurement of AGD in humans has been discussed in the literature, to our knowledge it has been measured formally in only two descriptive studies of females. Because AGD has been an easy-to-measure, sensitive outcome in animals studies, we developed and implemented an anthropometric protocol for measurement of AGD in human males as well as females. METHODS: We first evaluated the reliability of the AGD measures in 20 subjects. Then measurements were taken on an additional 87 newborns (42 females, 45 males). All subjects were from Morelos, Mexico. RESULTS: The reliability (Pearson r) of the AGD measure was, for females 0.50, and for males, 0.64. The between-subject variation in AGD, however, was much greater than the variation due to measurement error. The AGD measure was about two-fold greater in males (mean, 22 mm) than in females (mean, 11 mm), and there was little overlap in the distributions for males and females. CONCLUSION: The sexual dimorphism of AGD in humans comprises prima facie evidence that this outcome may respond to in utero exposure to hormonally active agents

Both mitochondrial DNA and mitonuclear gene mutations cause hearing loss through cochlear dysfunction.

PFC is a Wellcome Trust Senior Fellow in Clinical Science (101876/Z/13/Z), and a UK NIHR Senior Investigator, who receives support from the Medical Research Council Mitochondrial Biology Unit (MC_UP_1501/2), the Wellcome Trust Centre for Mitochondrial Research (096919Z/11/Z), the Medical Research Council (UK) Centre for Translational Muscle Disease research (G0601943), EU FP7 TIRCON, and the National Institute for Health Research (NIHR) Biomedical Research Centre based at Cambridge University Hospitals NHS Foundation Trust and the University of Cambridge. PJK is a Wellcome Trust Clinical research fellow (101700/Z/13/Z). PYWM is supported by a Clinician Scientist Fellowship Award (G1002570) from the Medical Research Council (UK), and also receives funding from Fight for Sight (UK) and the UK National Institute of Health Research (NIHR) as part of the Rare Diseases Translational Research Collaboration. The views expressed are those of the author(s) and not necessarily those of the NHS, the NIHR or the Department of Health. This study received support from the MRC Mitochondrial Disease Patient Cohort (http://mitocohort.ncl.ac.uk/).This is the final version of the article. It first appeared from Oxford University Press via https://doi.org/10.1093/brain/aww05

The DEAD-box RNA Helicase DDX6 is Required for Efficient Encapsidation of a Retroviral Genome

Viruses have to encapsidate their own genomes during the assembly process. For most RNA viruses, there are sequences within the viral RNA and virion proteins needed for high efficiency of genome encapsidation. However, the roles of host proteins in this process are not understood. Here we find that the cellular DEAD-box RNA helicase DDX6 is required for efficient genome packaging of foamy virus, a spumaretrovirus. After infection, a significant amount of DDX6, normally concentrated in P bodies and stress granules, re-localizes to the pericentriolar site where viral RNAs and Gag capsid proteins are concentrated and capsids are assembled. Knockdown of DDX6 by siRNA leads to a decreased level of viral nucleic acids in extracellular particles, although viral protein expression, capsid assembly and release, and accumulation of viral RNA and Gag protein at the assembly site are little affected. DDX6 does not interact stably with Gag proteins nor is it incorporated into particles. However, we find that the ATPase/helicase motif of DDX6 is essential for viral replication. This suggests that the ATP hydrolysis and/or the RNA unwinding activities of DDX6 function in moderating the viral RNA conformation and/or viral RNA-Gag ribonucleoprotein complex in a transient manner to facilitate incorporation of the viral RNA into particles. These results reveal a unique role for a highly conserved cellular protein of RNA metabolism in specifically re-locating to the site of viral assembly for its function as a catalyst in retroviral RNA packaging