Silac mouse for quantitative proteomics uncovers kindlin-3 as an essential factor for red blood cell function

Stable isotope labeling by amino acids in cell culture (SILAC) has become a versatile tool for quantitative, mass spectrometry (MS)-based proteomics. Here, we completely label mice with a diet containing either the natural or the 13C6-substituted version of lysine. Mice were labeled over four generations with the heavy diet, and development, growth, and behavior were not affected. MS analysis of incorporation levels allowed for the determination of incorporation rates of proteins from blood cells and organs. The F2 generation was completely labeled in all organs tested. SILAC analysis from various organs lacking expression of β1 integrin, β-Parvin, or the integrin tail-binding protein Kindlin-3 confirmed their absence and disclosed a structural defect of the red blood cell membrane skeleton in Kindlin-3-deficient erythrocytes. The SILAC-mouse approach is a versatile tool by which to quantitatively compare proteomes from knockout mice and thereby determine protein functions under complex in vivo conditions

Scalable production of large quantities of defect-free few-layer graphene by shear exfoliation in liquids

To progress from the laboratory to commercial applications, it will be necessary to develop industrially scalable methods to produce large quantities of defect-free graphene. Here we show that high-shear mixing of graphite in suitable stabilizing liquids results in large-scale exfoliation to give dispersions of graphene nanosheets. X-ray photoelectron spectroscopy and Raman spectroscopy show the exfoliated flakes to be unoxidized and free of basal-plane defects. We have developed a simple model that shows exfoliation to occur once the local shear rate exceeds 10(4) s(-1). By fully characterizing the scaling behaviour of the graphene production rate, we show that exfoliation can be achieved in liquid volumes from hundreds of millilitres up to hundreds of litres and beyond. The graphene produced by this method performs well in applications from composites to conductive coatings. This method can be applied to exfoliate BN, MoS2 and a range of other layered crystals

Yielding and flow of highly concentrated, few-layer graphene suspensions

For a wide range of applications of graphene suspensions, a thorough understanding of their rheological properties is crucial. We probe the microstructure of dense suspensions of micron-sized, few-layer, defect-free graphene platelets by measuring their viscoelastic properties at various concentrations up to 39 mg ml-1. We propose a model to relate the yield strain to the mesh size of the microstructure as a function of volume fraction ?. From the yield stress measurements we infer the typical bond energy (?20 kBT) and ? dependence of the bond number density. These results allow us to express the steady shear viscosity for Peclet number Pe < 10 in terms of the platelet dimensions, bond energy and ? using a relaxation ansatz

Analysis of the foam-forming of non-woven lightweight fibrous materials using X-ray tomography

Foam-forming has in the past predominantly been used to create two-dimensional sheet-like fibrous materials. Allowing the foam to drain freely and decay under gravity, rather than applying a vacuum to remove it rapidly, we can produce lightweight three-dimensional fibrous structures from cellulose fibres, of potential use for thermal and acoustic insulation. ?muC scanning of the fibrous materials enable us to determine both void size distributions and also distributions of fibre orientations. Through image analysis and uniaxial compression testing, we find that the orientation of the fibres, rather than the size of the voids, determine the compressive strength of the material. The fibrous samples display a layering of the fibres perpendicular to the direction of drainage of the precursor liquid foam. This leads to an anisotropy of the compressive behaviour of the samples. Varying the initial liquid fraction of the foam allows for tuning of the compressive strength. We show an increase in over seven times can be achieved for samples of the same density (13 kg.m-3)

2D foams above the jamming transition: Deformation matters

Jammed soft matter systems are often modelled as dense packings of overlapping soft spheres, thus ignoring particle deformation. For 2D (and 3D) soft disks packings, close to the critical packing fraction ?c, this results in an increase of the average contact number Z with a square root in ? ? ?c. Using the program PLAT, we find that in the case of idealised two-dimensional foams, close to the wet limit, Z increases linearly with ? ? ?c, where ? is the gas fraction. This result is consistent with the different distributions of separations for soft disks and foams at the critical packing fraction. Thus, 2D foams close to the wet limit are not well described as random packings of soft disks, since bubbles in a foam are deformable and adjust their shape. This is not captured by overlapping circular disks

Stabilizing effect of a magnetic &#64257;eld on a gas bubble produced at a microelectrode

The effect on the growth of a single hydrogen bubble of a magnetic field oriented normal to the surface of a microelectrode is investigated. Overpotential oscillations due to periodic growth and detachment of bubbles are correlated with high-speed camera images and particle tracking was used to follow the flow pattern. The bubbles on both horizontal and vertical microelectrodes grow bigger in the field, doubling in diameter before they break away in 5 tesla. The effect is related to the influence of the magnetic field on the breakoff mechanism rather than to Ekman pumping driven by the Lorentz force

Fiber Reinforced Cartilage ECM Functionalized Bioinks for Functional Cartilage Tissue Engineering

Focal articular cartilage (AC) defects, if left untreated, can lead to debilitating diseases such as osteoarthritis. While several tissue engineering strategies have been developed to promote cartilage regeneration, it is still challenging to generate functional AC capable of sustaining high load-bearing environments. We developed a new class of cartilage extracellular matrix (cECM)-functionalized alginate bioink for the bioprinting of cartilaginous tissues. The bioinks were 3D-printable, supported mesenchymal stem cell (MSC) viability post-printing and robust chondrogenesis in vitro, with the highest levels of COLLII and ACAN expression observed in bioinks containing the highest concentration of cECM. Enhanced chondrogenesis in cECM-functionalized bioinks was also associated with progression along an endochondral-like pathway, as evident by increases in RUNX2 expression and calcium deposition in vitro. The bioinks loaded with MSCs and TGF-?3 were also found capable of supporting robust chondrogenesis, opening the possibility of using such bioinks for direct ?print-and-implant? cartilage repair strategies. Finally, we demonstrated that networks of 3D-printed polycaprolactone fibers with compressive modulus comparable to native AC could be used to mechanically reinforce these bioinks, with no loss in cell viability. It is envisioned that combinations of such biomaterials can be used in multiple-tool biofabrication strategies for the bioprinting of biomimetic cartilaginous implants

Statistical integration of multi-omics and drug screening data from cell lines.

Data integration methods are used to obtain a unified summary of multiple datasets. For multi-modal data, we propose a computational workflow to jointly analyze datasets from cell lines. The workflow comprises a novel probabilistic data integration method, named POPLS-DA, for multi-omics data. The workflow is motivated by a study on synucleinopathies where transcriptomics, proteomics, and drug screening data are measured in affected LUHMES cell lines and controls. The aim is to highlight potentially druggable pathways and genes involved in synucleinopathies. First, POPLS-DA is used to prioritize genes and proteins that best distinguish cases and controls. For these genes, an integrated interaction network is constructed where the drug screen data is incorporated to highlight druggable genes and pathways in the network. Finally, functional enrichment analyses are performed to identify clusters of synaptic and lysosome-related genes and proteins targeted by the protective drugs. POPLS-DA is compared to other single- and multi-omics approaches. We found that HSPA5, a member of the heat shock protein 70 family, was one of the most targeted genes by the validated drugs, in particular by AT1-blockers. HSPA5 and AT1-blockers have been previously linked to α-synuclein pathology and Parkinson's disease, showing the relevance of our findings. Our computational workflow identified new directions for therapeutic targets for synucleinopathies. POPLS-DA provided a larger interpretable gene set than other single- and multi-omic approaches. An implementation based on R and markdown is freely available online