Investigating cellular signaling reactions in single attoliter vesicles

Understanding cellular signaling mediated by cell surface receptors is key to modern biomedical research and drug development. The discovery of a growing number of potential molecular targets and therapeutic compounds requires downscaling and accelerated functional screening. Receptor-mediated cellular responses are typically investigated on single cells or cell populations. Here, we show how to monitor cellular signaling reactions at a yet unreached miniaturization level. On the basis of our observations, cytochalasin induces mammalian cells to extrude from their plasma membrane submicrometer-sized native vesicles. They comprise functional cell surface receptors correctly exposing their extracellular ligand binding sites on the outer vesicle surface and retaining cytosolic proteins in the vesicle interior. As a prototypical example, ligand binding to the ionotropic 5-HT(3) receptor and subsequent transmembrane Ca(2+) signaling were monitored in single attoliter vesicles. Thus, native vesicles are the smallest autonomous containers capable of performing cellular signaling reactions under physiological conditions. Because a single cell delivers about 50 native vesicles, which can be isolated and addressed as individuals, our concept allows multiple functional analyses of individual cells having a limited availability and opens new vistas for miniaturized bioanalytics.     

Determination of 226Ra in solid samples of few milligrams after mineralisation and measurement by solid scintillation

A protocol is proposed for the measurement of ²²⁶Ra on 150 mg of solid sample without radiochemistry. To evaluate the performance of this method, standard samples were used and the results were in good agreement with reference values. The detection limit obtained is about 130 Bq kg⁻¹ (dry weight) without mineralised solution concentration by evaporation. A concentration of the solution by 4 and/or an increase of sample mass by 4 in the case of microwave digestion system, allows achieving a detection limit of 30 Bq kg⁻¹ (dry weight) and thus measuring ²²⁶Ra in most soils. This method could also be used for NORM sites on soil and sediment samples.

     

Neighborhood unions and extremal spanning trees

We generalize a known sufficient condition for the traceability of a graph to a condition for the existence of a spanning tree with a bounded number of leaves. Both of the conditions involve neighborhood unions. Further, we present two results on spanning spiders (trees with a single branching vertex). We pose a number of open questions concerning extremal spanning trees.     

UV-ABC screens of luteolin derivatives compared to edelweiss extract

Pure luteolin is a remarkably heat (200°C/6 days) and UV stable UV-A screen, however, native luteolin enriched to 37% in an edelweiss extract lost its UV-A screen properties upon UV irradiation (～4MJm(-2)). This contrasting behavior led to the examination of a series of purified luteolin derivatives as UV screen candidates. 3',4',5,7-Tetralipoyloxyflavones were synthesized from luteolin (3',4',5,7-tetrahydroxyflavone) and fatty acid chlorides. These acylated semi-biomolecules show a hypsochromic shift in UV-Vis spectra of about Δλ(A→B)=58nm and absorbed in the centre of the harmful UV-B band (λ(max)=295nm). Luteolin was also hydroxyethylated with Br(CH(2))(2)OH. This substitution has no effect on the λ(max)=330nm absorption of luteolin (UV-A band). Finally the natural 4'-O-β-glucosyl-3',5,7-trihydroxyflavone was extracted from edelweiss and used as a purified natural benchmark. Glycosylated and hydroxyethylated luteolin are both UV stable. Fully acylated luteolin derivatives degrade upon UV exposure to a stable UV-C screen with a hypsochroic shift Δλ(B→C)=35nm. All in all, three molecular structures based on luteolin with sunscreen properties were found, distinguishable in: UV-A, UV-B, and UV-C filters. The natural product based UV-absorbers show promise as alternatives to synthetic molecules and nanoparticles in sunscreen products.     

Concise synthesis of enantiopure (S)- and (R)-α-trifluoromethyl aspartic acid and α-trifluoromethyl serine from chiral trifluoromethyl oxazolidines (Fox) via a Strecker-type reaction

A concise synthesis of enantiopure (S)- and (R)-α-Tfm-aspartic acid and α-Tfm-serine is reported. The key step involves a Strecker-type reaction on chiral CF₃-oxazolidines (Fox) derived from ethyl 4,4,4-trifluoroacetoacetate (ETFAA) or ethyl trifluoropyruvate.     

Highly diastereoselective synthesis of enantiopure β-trifluoromethyl β-amino alcohols from chiral trifluoromethyl oxazolidines (Fox)

The organolithium species addition to 2-hydroxymethyl fluorinated oxazolidines (Fox) provides a highly diastereoselective and straightforward route for the synthesis of enantiopure trifluoromethyl β-amino alcohols quaternarized at the β-position.     

Computer simulation of the rheology of concentrated star polymer suspensions

We use particle-based computer simulations to study the rheology of suspensions of high-functionality star polymers with long entangled arms. Such particles have properties which are intermediate between those of soft colloidal particles and entangled polymer chains. In the simulations, each star polymer is coarse-grained to a single particle. In order to faithfully reproduce dynamical properties, it is very important to not only include time-averaged interactions (potentials of mean force) but to also account for transient interactions induced by entanglements between the arms of different star polymers. Using a model which has all these features, it is found that, for sufficiently high shear rates, the start-up shear stress displays an overshoot. With increasing concentration, the core interactions increasingly dominate the initial stress response, leading to a maximum in the stress overshoot at relatively low strain values (0.1 to 0.5). Transient forces start to dominate after this initial stage. In a simulated experiment in which the shear rate is suddenly stepped-down from a high to a lower value, the stress shows a clear undershoot, with the minimum stress again at a relatively low strain value (based on the new shear rate). Finally, it is shown that a stress plateau develops in the flow curve. This plateau is absent when the transient forces between the polymer stars are not taken into account.     

Collision of Almost Parallel Vortex Filaments

We investigate the occurrence of collisions in the evolution of vortex filaments through a system introduced by Klein, Majda, and Damodaran and Zakharov . We first establish rigorously the existence of a pair of almost parallel vortex filaments, with opposite circulation, colliding at some point in finite time. The collision mechanism is based on one of the self‐similar solutions of the model, described by the authors in an earlier work. In the second part of this paper we extend this construction to the case of an arbitrary number of filaments, with polygonal symmetry, that are perturbations of a configuration of parallel vortex filaments forming a polygon, with or without its center, rotating with constant angular velocity.© 2016 Wiley Periodicals, Inc.