Flow Cytometry Analysis of Circulating Extracellular Vesicle Subtypes from Fresh Peripheral Blood Samples

Extracellular vesicles (EVs) are released by shedding during different physiological processes and are increasingly thought to be new potential biomarkers. However, the impact of pre-analytical processing phases on the final measurement is not predictable and for this reason, the translation of basic research into clinical practice has been precluded. Here we have optimized a simple procedure in combination with polychromatic flow cytometry (PFC), to identify, classify, enumerate, and separate circulating EVs from different cell origins. This protocol takes advantage of a lipophilic cationic dye (LCD) able to probe EVs. Moreover, the application of the newly optimized PFC protocol here described allowed the obtainment of repeatable EVs counts. The translation of this PFC protocol to fluorescence-activated cell sorting allowed us to separate EVs from fresh peripheral blood samples. Sorted EVs preparations resulted particularly suitable for proteomic analyses, which we applied to study their protein cargo. Here we show that LCD staining allowed PFC detection and sorting of EVs from fresh body fluids, avoiding pre-analytical steps of enrichment that could impact final results. Therefore, LCD staining is an essential step towards the assessment of EVs clinical significance.     

Remapping of Adult-Born Neuron Activity during Fear Memory Consolidation in Mice

The mammalian hippocampal dentate gyrus is a unique memory circuit in which a subset of neurons is continuously generated throughout the lifespan. Previous studies have shown that the dentate gyrus neuronal population can hold fear memory traces (i.e., engrams) and that adult-born neurons (ABNs) support this process. However, it is unclear whether ABNs themselves hold fear memory traces. Therefore, we analyzed ABN activity at a population level across a fear conditioning paradigm. We found that fear learning did not recruit a distinct ABN population. In sharp contrast, a completely different ABN population was recruited during fear memory retrieval. We further provide evidence that ABN population activity remaps over time during the consolidation period. These results suggest that ABNs support the establishment of a fear memory trace in a different manner to directly holding the memory. Moreover, this activity remapping process in ABNs may support the segregation of memories formed at different times. These results provide new insight into the role of adult neurogenesis in the mammalian memory system.     

Machine learning in corporate credit rating assessment using the expanded audit report

Machine Learning - We investigate whether key audit matter (KAM) paragraphs disclosed in extended audit reports—paragraphs in which the auditor highlights significant risks and critical...     

Reversible Thermochromic Polymeric Thin Films Made of Ultrathin 2D Crystals of Coordination Polymers Based on Copper(I)‐Thiophenolates

A one‐pot reaction between Cu(BF₄)₂·xH₂O and 4‐mercaptobenzoic acid in acetone or methanol gives rise to the formation of lamellar microcrystals of two Cu(I)‐thiophenolate‐based coordination polymers (CPs) with the formulas [CuCT] _n ( 1 ) (CT = 4‐carboxy‐thiophenolate) and [CuMCT]_n ( 2 ) (MCT = 4‐methoxycarbonyl‐thiophenolate). Both 1 and 2 show a reversible luminescent thermochromic behavior upon cooling, changing their color from pale yellow to green to orange in the case of 1 , and from pale orange to green in the case of 2 . It is shown that the lamellar character of these crystals, which exhibit micrometer lateral dimensions and sub‐micrometer/nanometer thicknesses, allows processing them with an organic polymer such as polyvinylidene difluoride (PVDF) to form thermochromic 1 @PVDF and 2 @PVDF thin films. These thermal stimuli‐responsive thin films are freestanding, free of macroscopic defects, and robust under mechanical bending stress, opening up the possibility to use them in, for example, 2D imaging sensor films.     

Synthesis,structural characterization and luminescent properties of Tb3+-doped AgLaP2O7 phosphors

Silver lanthanum diphosphates doped with terbium, AgLa_1-xTb_xP₂O₇ (x=0%, 1%, 5%, 10%, 15% and 20%), were prepared by solid state reaction at T=500°. The obtained compounds were investigated by means of a multi-methodological approach, involving the Scanning Electron Microscopy (SEM) equipped with energy dispersive X-Ray spectroscopy (EDS) for morphological investigation and semi-quantitative chemical analysis, respectively; powder X-ray diffraction (PXRD) for structural characterization and Rietveld refinement; Fourier Transform InfraRed (FTIR) and Raman spectroscopies for qualitative study. Finally, emission spectra were collected in order to detect the fluorescence properties of the compounds.The unit cell parameters and the space group of all the family members were determined by PXRD data. The compounds crystallize in the orthorhombic Pnma space group, with the AgLaP₂O₇ cell constants equal to: a?=?8.6706(1) Å, b?=?5.3218(1) Å, c?=?12.8839(1) Å and cell volume V?=?594.51(1) ų. A decreasing trend for unit cell parameters was observed at increase of dopant concentration. For the pure phase, the investigation was completed with the structure solution via Direct Methods and Rietveld refinement. The crystal structure consists of compact layers of LaO₉ polyhedra, down the c axis, bridged by P₂O₇ diphosphate groups and by zig-zag chains of AgO₉ polyhedra down b.The FTIR and Raman analysis supports the chemical structure, highlighting the deformation (δPO₃) and stretching (νPO₃) vibration of PO₃ groups, and the characteristic bands of P₂O₇^4- groups attributed to the symmetric and asymmetric stretching (ν_s and ν_as) of P-O-P bridge, respectively.The luminescence properties of Tb³⁺ activating ion in AgLa_1-xTb_xP₂O₇ (x=1%, 5%, 10%, 15% and 20%) were also examined. No concentration quenching of the main ⁵D₄-→⁷F₅ emission, responsible for the well known green emission of Tb³⁺ ion, was observed in the studied concentration range, whereas a concentration quenching of the emission from ⁵D₃ level, due to ⁵D₃^-→⁵D₄ cross relaxation process, was revealed.     

Electrospray‐assisted fabrication of core‐shell arabinoxylan gel particles for insulin and probiotics entrapment

This work presents the fabrication and characterization of electro‐sprayed core‐shell particles that were composed of maize bran arabinoxylans (MBAX) with insulin in the core, and maize wastewater arabinoxylans (MWAX) with Bifidobacterium in the shell. Two concentrations of MBAX (3% and 6% w/v) and MWAX (6% and 10% w/v) were evaluated. The particles fabricated with MBAX at 6% (w/v) in the core and MWAX at 10% (w/v) in the shell were more stable, presented spherical shape and no aggregation being therefore selected to be loaded with insulin and probiotics. These particles presented a size of 2.9 mm. Scanning electron microscopy analysis of the particle cross section revealed the presence of both, a smooth (shell) and a porous (core) microstructure. Confocal laser scanning microscopy confirmed the core‐shell structure of the particles and the viability of the probiotic entrapped. Gastrointestinal simulation strongly suggests that these particles are not degraded in the stomach and small intestine and that 76% of the carried insulin is released in colon. These results indicate that insulin and Bifidobacterium encapsulation by tetraaxial electro spraying can be a feasible and adequate technique to produce arabinoxylan capsules containing both insulin and probiotics. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2018 , 135, 46411.     

Study of the hot-wire TIG process with AISI-316L filler material,analysing the effect of magnetic arc blow on the dilution of the weld bead

The TIG process with hot-wire addition is described as a highly productive process in comparison with conventional TIG process with cold-wire and achieves large deposition rates and low dilution percentages, characteristics that permit to the process to be an optimum alternative for cladding applications. There are many options of electrical circuit configurations to heat the addition wire in the TIG Hot-Wire process, highlighting the utilization of the continuous pulsed current and the alternating current. In the present paper were studied two configurations of the electrical circuit for heating the addition materials, which in theory provoke different behaviours in the welding arc, and therefore, in the geometrical properties of the welding bead too. A first analysis done on the welding arc, which demonstrates that to heat the wire with continuous constant current generates a permanent magnetic blow over the arc. According to trials made, the direction and intensity of the magnetic blow will depend on the polarity and on the current value to heat the wire. A macrographic analysis of the weld beads demonstrated that the best results were for the tests made with hot-wire, achieving dilutions up to 2% for a wire velocity of 7.5 m/min, 1 kW of power for to heat the wire and using a welding velocity of 30 cm/min.     

Overview of the Therapeutic Potential of Aptamers Targeting Coagulation Factors

Aptamers are single-stranded DNA or RNA sequences that bind target molecules with high specificity and affinity. Aptamers exhibit several notable advantages over protein-based therapeutics. Aptamers are non-immunogenic, easier to synthesize and modify, and can bind targets with greater affinity. Due to these benefits, aptamers are considered a promising therapeutic candidate to treat various conditions, including hematological disorders and cancer. An active area of research involves developing aptamers to target blood coagulation factors. These aptamers have the potential to treat cardiovascular diseases, blood disorders, and cancers. Although no aptamers targeting blood coagulation factors have been approved for clinical use, several aptamers have been evaluated in clinical trials and many more have demonstrated encouraging preclinical results. This review summarized our knowledge of the aptamers targeting proteins involved in coagulation, anticoagulation, fibrinolysis, their extensive applications as therapeutics and diagnostics tools, and the challenges they face for advancing to clinical use.