Intravital FRET: Probing Cellular and Tissue Function in Vivo

The development of intravital Förster Resonance Energy Transfer (FRET) is required to probe cellular and tissue function in the natural context: the living organism. Only in this way can biomedicine truly comprehend pathogenesis and develop effective therapeutic strategies. Here we demonstrate and discuss the advantages and pitfalls of two strategies to quantify FRET in vivo—ratiometrically and time-resolved by fluorescence lifetime imaging—and show their concrete application in the context of neuroinflammation in adult mice.     

Modulators of Macrophage Polarization Influence Healing of the Infarcted Myocardium

To diminish heart failure development after acute myocardial infarction (AMI), several preclinical studies have focused on influencing the inflammatory processes in the healing response post-AMI. The initial purpose of this healing response is to clear cell debris of the injured cardiac tissue and to eventually resolve inflammation and support scar tissue formation. This is a well-balanced reaction. However, excess inflammation can lead to infarct expansion, adverse ventricular remodeling and thereby propagate heart failure development. Different macrophage subtypes are centrally involved in both the promotion and resolution phase of inflammation. Modulation of macrophage subset polarization has been described to greatly affect the quality and outcome of healing after AMI. Therefore, it is of great interest to reveal the process of macrophage polarization to support the development of therapeutic targets. The current review summarizes (pre)clinical studies that demonstrate essential molecules involved in macrophage polarization that can be modulated and influence cardiac healing after AMI.     

The calculation of the heat release rate by oxygen consumption in a controlled‐atmosphere cone calorimeter

The standard cone calorimeter according to ASTM E 1354 and ISO 5660 enables reaction‐to‐fire tests to be performed in ambient atmospheric conditions. A controlled‐atmosphere chamber modifies the standard apparatus in a way that allows tests to be performed in nonambient conditions as well. The enclosed chamber is placed underneath the standard exhaust hood and does not have a closed connection to the hood. With this open arrangement, the exhaust gases are diluted by excess air drawn in from the laboratory surroundings. Heat‐induced changes in the consequential dilution ratio affect the calculation of fire quantities and, when neglected, lead to deviations of up to 30% in heat release rate. The paper introduces a test protocol and equations to calculate the heat release rate taking dilution effects into account. A mathematical correction is shown that compensates for the dilution effects while avoiding extensive mechanical changes in the equipment. Copyright © 2013 John Wiley & Sons, Ltd.     

Engineering egress data considering pedestrians with reduced mobility

To quantify the evacuation process, evacuation practitioners use engineering egress data describing the occupant movement characteristics. These data are typically based to young and fit populations. However, the movement abilities of occupants who might be involved in evacuations are becoming more variable—with the building populations of today typically including increasing numbers of individuals: with impairments or who are otherwise elderly or generally less mobile. Thus, there will be an increasing proportion of building occupants with reduced ability to egress. For safe evacuation, there is therefore a need to provide valid engineering egress data considering pedestrians with disabilities. Gwynne and Boyce recently compiled a series of data sets related to the evacuation process to support practitioner activities in the chapter Engineering Data in the SFPE Handbook of Fire Protection Engineering. This paper supplements these data sets by providing information on and presenting data obtained from additional research related to the premovement and horizontal movement of participants with physical‐, cognitive‐, or age‐related disabilities. The aim is to provide an overview of currently available data sets related to, and key factors affecting the egress performance of, mixed ability populations which could be used to guide fire safety engineering decisions in the context of building design.     

Interlaboratory study on lipid oxidation during accelerated storage trials with rapeseed and sunflower oil analyzed by conjugated dienes as primary oxidation products

Accelerated storage tests are frequently used to assess the oxidative stability of foods and related systems due to its reproducibility. Various methods and experimental conditions are used to measure lipid oxidation. Differences between laboratories make it necessary to determine the repeatability and reproducibility of oxidation tests performed under the same conditions. The objective of the present interlaboratory study was to evaluate the outcome of a storage test for two different bulk oils, sunflower oil (SFO) and rapeseed oil (RSO), during a period of 9 weeks at 20°C, 30°C, 40°C, and 60°C. Sixteen laboratories were provided with bottled oils and conducted the storage tests according to a detailed protocol. Lipid oxidation was monitored by the formation of conjugated dienes (CD) and the activation energy (E_a) was determined for comparative purposes and statistically evaluated. An increase in CD formation was observed for both oils when the storage temperature was increased in all laboratories. The E_a,1 ranged from 47.9 to 73.3 kJ mol⁻¹ in RSO and from 27.8 to 62.6 kJ mol⁻¹ in SFO, with average values of 58.2 and 46.8 kJ mol⁻¹, respectively. The reproducibility coefficients were 10.9% and 18.2% for RSO and SFO, respectively. Practical applications: In order to compare results on oxidative stability of foods derived from different studies, the reproducibility of storage tests and methods employed to evaluate the oxidation level should be considered. This study provides fundamental data on the reproducibility of lipid oxidation under accelerated storage conditions and defines important parameters to be considered for the conduction of experiments.     

Optical Properties of Metacrystals Based on Protein Nanocages

The optical properties of 3D metacrystals made of gold nanoparticles in protein nanocages are studied. These metacrystals have sizes of tens of micrometers and are of high structural and optical quality. Through microspectroscopy measurements and model calculations it is demonstrated that the metacrystals show plasmonic absorption in the green wavelength range and are largely transparent in the red and infrared ranges. By using empty nanocages as placeholders in the metacrystal lattice, it is possible to control how strongly the metamaterial absorbs. Measurements on a pyramidal metacrystal show that it deflects visible light. The deflection shows evidence for anomalous refraction at short wavelengths and normal refraction at long wavelengths. The refractive dispersion is ascribed to the optical dispersion relation of the plasmonic metamaterial.     

Common Muscle Metabolic Signatures Highlight Arginine and Lysine Metabolism as Potential Therapeutic Targets to Combat Unhealthy Aging

Biological aging research is expected to reveal modifiable molecular mechanisms that can be harnessed to slow or possibly reverse unhealthy trajectories. However, there is first an urgent need to define consensus molecular markers of healthy and unhealthy aging. Established aging hallmarks are all linked to metabolism, and a ‘rewired’ metabolic circuitry has been shown to accelerate or delay biological aging. To identify metabolic signatures distinguishing healthy from unhealthy aging trajectories, we performed nontargeted metabolomics on skeletal muscles from 2-month-old and 21-month-old mice, and after dietary and lifestyle interventions known to impact biological aging. We hypothesized that common metabolic signatures would highlight specific pathways and processes promoting healthy aging, while revealing the molecular underpinnings of unhealthy aging. Here, we report 50 metabolites that commonly distinguished aging trajectories in all cohorts, including 18 commonly reduced under unhealthy aging and 32 increased. We stratified these metabolites according to known relationships with various aging hallmarks and found the greatest associations with oxidative stress and nutrient sensing. Collectively, our data suggest interventions aimed at maintaining skeletal muscle arginine and lysine may be useful therapeutic strategies to minimize biological aging and maintain skeletal muscle health, function, and regenerative capacity in old age.     

Proton-Detected Solid-State NMR of the Cell-Free Synthesized α-Helical Transmembrane Protein NS4B from Hepatitis C Virus

Proton-detected 100 kHz magic-angle-spinning (MAS) solid-state NMR is an emerging analysis method for proteins with only hundreds of microgram quantities, and thus allows structural investigation of eukaryotic membrane proteins. This is the case for the cell-free synthesized hepatitis C virus (HCV) nonstructural membrane protein 4B (NS4B). We demonstrate NS4B sample optimization using fast reconstitution schemes that enable lipid-environment screening directly by NMR. 2D spectra and relaxation properties guide the choice of the best sample preparation to record 2D ¹H-detected ¹H,¹⁵N and 3D ¹H,¹³C,¹⁵N correlation experiments with linewidths and sensitivity suitable to initiate sequential assignments. Amino-acid-selectively labeled NS4B can be readily obtained using cell-free synthesis, opening the door to combinatorial labeling approaches which should enable structural studies.     

Chemotactic and Angiogenic Potential of Mineralized Collagen Scaffolds Functionalized with Naturally Occurring Bioactive Factor Mixtures to Stimulate Bone Regeneration

To develop cost-effective and efficient bone substitutes for improved regeneration of bone defects, heparin-modified mineralized collagen scaffolds were functionalized with concentrated, naturally occurring bioactive factor mixtures derived from adipose tissue, platelet-rich plasma and conditioned medium from a hypoxia-treated human bone marrow-derived mesenchymal stem cell line. Besides the analysis of the release kinetics of functionalized scaffolds, the bioactivity of the released bioactive factors was tested with regard to chemotaxis and angiogenic tube formation. Additionally, functionalized scaffolds were seeded with human bone marrow-derived mesenchymal stromal cells (hBM-MSC) and their osteogenic and angiogenic potential was investigated. The release of bioactive factors from the scaffolds was highest within the first 3 days. Bioactivity of the released factors could be confirmed for all bioactive factor mixtures by successful chemoattraction of hBM-MSC in a transwell assay as well as by the formation of prevascular structures in a 2D co-culture system of hBM-MSC and human umbilical vein endothelial cells. The cells seeded directly onto the functionalized scaffolds were able to express osteogenic markers and form tubular networks. In conclusion, heparin-modified mineralized collagen scaffolds could be successfully functionalized with naturally occurring bioactive factor mixtures promoting cell migration and vascularization.