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Store-operated [Ca.sup.2+] entry (SOCE) is the major route of [Ca.sup.2+] influx in platelets. The [Ca.sup.2+] sensor stromal interaction molecule 1 (STIM1) triggers SOCE by forming punctate structures with the [Ca.sup.2+] channel Orai1 and the inositol trisphosphate receptor ([IP.sub.3]R), thereby linking the endo-/sarcoplasmic reticulum to the plasma membrane. Here, we identified the BAR domain superfamily member bridging integrator 2 (BIN2) as an interaction partner of STIM1 and [IP.sub.3]R in platelets. Deletion of platelet BIN2 ([Bin2.sup.fl/fl,Pf4,Cre] mice) resulted in reduced [Ca.sup.2+] store release and [Ca.sup.2+] influx in response to all tested platelet agonists. These defects were a consequence of impaired [IP.sub.3]R function in combination with defective STIM1-mediated SOC channel activation, while [Ca.sup.2+] store content and agonist-induced [IP.sub.3] production were unaltered. This severely defective [Ca.sup.2+] signaling translated into impaired thrombus formation under flow and a protection of [Bin2.sup.fl/fl,Pf4,Cre] mice in models of arterial thrombosis and stroke. Our results establish BIN2 as a central regulator of platelet activation in thrombosis and thrombo- inflammatory disease settings., Introduction Platelet activation and aggregation at the injured vessel wall is essential for primary hemostasis, but may also trigger occlusion of diseased vessels, resulting in acute ischemia and infarction of [...]

Xiphophorus fish exhibit a clear phenotypic polymorphism in puberty onset and reproductive strategies of males. In X. nigrensis and X. multilineatus, puberty onset is genetically determined and linked to a melanocortin 4 receptor (Mc4r) polymorphism of wild-type and mutant alleles on the sex chromosomes. We hypothesized that Mc4r mutant alleles act on wild-type alleles by a dominant negative effect through receptor dimerization, leading to differential intracellular signaling and effector gene activation. Depending on signaling strength, the onset of puberty either occurs early or is delayed. Here, we show by Förster Resonance Energy Transfer (FRET) that wild-type Xiphophorus Mc4r monomers can form homodimers, but also heterodimers with mutant receptors resulting in compromised signaling which explains the reduced Mc4r signaling in large males. Thus, hetero- vs. homo- dimerization seems to be the key molecular mechanism for the polymorphism in puberty onset and body size in male fish. [ABSTRACT FROM AUTHOR]

G-protein-coupled receptors (GPCRs) are hypothesized to possess molecular mobility over a wide temporal range. Until now the temporal range has not been fully accessible due to the crucially limited temporal range of available methods. This in turn, may lead relevant dynamic constants to remain masked. Here, we expand this dynamic range by combining fluorescent techniques using a spot confocal setup. We decipher mobility constants of β2-adrenergic receptor over a wide time range (nanosecond to second). Particularly, a translational mobility (10 µm²/s), one order of magnitude faster than membrane associated lateral mobility that explains membrane protein turnover and suggests a wider picture of the GPCR availability on the plasma membrane. And a so far elusive rotational mobility (1-200 µs) which depicts a previously overlooked dynamic component that, despite all complexity, behaves largely as predicted by the Saffman-Delbrück model. The mobility of the β2-adrenergic receptor, from the nanosecond to the second range, is revealed by combining several fluorescent spectroscopy techniques. These data also show a previously hidden mobility of this GPCR. [ABSTRACT FROM AUTHOR]

All hematopoietic cells that develop in the bone marrow must cross the endothelial barrier to enter the blood circulation. Blood platelets, however, are released by bigger protrusions of huge progenitor cells, named megakaryocytes, and enter the blood stream as so-called proplatelets before fragmenting into mature platelets. Recently, a second function of megakaryocytes has been identified, as they modulate the quiescence of hematopoietic stem cells, mostly via different soluble factors. We know from light sheet fluorescence microscopy images that megakaryocytes are distributed throughout the bone marrow facing a dense vascular network. Here, we used such three-dimensional images to provide a realistic simulation template reflecting the in vivo cell-vessel distributions resulting in reliable whole-bone analysis in silico Combining this approach with an automated image analysis pipeline, we found that megakaryocytes influence migration of neutrophils and hematopoietic stem cells, and thus act as biomechanical restrainers modulating cell mobility and extravasation. Indeed, as a consequence of increased megakaryocyte volumes in platelet-depleted mice neutrophil mobility was reduced in these animals., (Copyright© 2020 Ferrata Storti Foundation.)

Madurahydroxylactone (MHL), a secondary metabolite with antibacterial activity was evaluated for its suitability to generate controlled drug release coatings on medical implant materials. A smooth and firmly attached layer could be produced from a precursor solution on various metallic implant materials. In physiological salt solutions these coatings dissolved within a time period up to one week. A combination of MHL with a broad spectrum fluoroquinolone antibiotic was used to create a coating that was active against all bacterial strains tested. The time period during which the coating remained active against Pseudomonas aeruginosa was investigated. The results indicated a delayed drug release from single layer coatings in the course of seven days. MHL was biocompatible in cell culture assays and could after a delay even serve as a cell adhesion substrate for human or murine cells. The findings indicate a potential for MHL for the generation of delayed release antimicrobial implant coatings.