Efficient organic monoliths prepared by γ-radiation induced polymerization in the evaluation of histone deacetylase inhibitors by capillary(nano)-high performance liquid chromatography and ion trap mass spectrometry

New monolithic HPLC columns were prepared by γ-radiation-triggered polymerization of hexyl methacrylate and ethylene glycol dimethacrylate monomers in the presence of porogenic solvents. Polymerization was carried out directly within capillary (250-200 μm I.D.) and nano (100-75 μm I.D.) fused-silica tubes yielding highly efficient columns for cap(nano)-LC applications. The columns were applied in the complete separation of core (H2A, H2B, H3, and H4) and linker (H1) histones under gradient elution with UV and/or electrospray ionization (ESI) ion trap mass spectrometry (MS) detections. Large selectivity towards H1, H2A-1, H2A-2, H2B, H3-1, H3-2 and H4 histones and complete separation were obtained within 8 min time windows, using fast gradients and very high linear flow velocities, up to 11 mm/s for high throughput applications. The method developed was the basis of a simple and efficient protocol for the evaluation of post-translational modifications (PTMs) of histones from NCI-H460 human non-small-cell lung cancer (NSCLC) and HCT-116 human colorectal carcinoma cells. The study was extended to monitoring the level of histone acetylation after inhibition of Histone DeACetylase (HDAC) enzymes with suberoylanilide hydroxamic acid (SAHA), the first HDAC inhibitor approved by the FDA for cancer therapy. Attractive features of our cap(nano)-LC/MS approach are the short analysis time, the minute amount of sample required to complete the whole procedure and the stability of the polymethacrylate-based columns. A lab-made software package ClustMass was ad hoc developed and used to elaborate deconvoluted mass spectral data (aligning, averaging, clustering) and calculate the potency of HDAC inhibitors, expressed through a Relative half maximal Inhibitory Concentration parameter, namely R_IC(50) and an averaged acetylation degree.     

Water evaporation from gel beads

Hydrogels are characterized by properties which make them ideal candidates for applications in several fields, such as drug delivery, biomedicine, and functional foods. Molecular diffusion out of a hydrogel matrix depends on their hydrodynamic radii and the mesh sizes within the matrix of the gel. A quantitative experimental and mathematical understanding of interactions, kinetics, and transport phenomena within complex hydrogel systems assists network design by identifying the key parameters and mechanisms that govern the rate and extent of solute release. In this article a calorimetric differential scanning calorimetry (DSC) study reports on the approach to parallel water effusion from a hydrogel matrix to the release of a model protein. The measurement of the water evaporation is taken as the simplest routine determination of a phenomenon that is basically due to a diffusive process through the porous structure of the gel and thermodynamically governed by the difference in the water chemical potential inside and outside of the bead. The analysis of the experimental calorimetric curves is made with the purpose of extracting several numerical parameters characteristic of each curve. The rationale is to develop a simple methodology to understand the release properties of the porous structure of the complex gel matrix by means of DSC.     

Optimization of a 2D photonic crystal add/drop multiplexer based on contra-directional coupling

We present a highly integrated add/drop multiplexer, where the contra-directional coupling is realized by phase matching two photonic crystal waveguides. The device band structure, the corresponding transmission and drop spectra, and the coupling length are carefully analysed. Different device configurations are discussed and by tailoring the coupling factor, we optimize the frequency response of the filter, obtaining a -sized channel selector, characterized by a very high drop efficiency.     

The palladium-phenanthroline catalyzed carbonylation of nitroarenes to diarylureas: Effect of chloride and diphenylphosphinic acid

The application of the palladium-phenanthroline catalytic system to the carbonylation of nitrobenzene in the presence of aniline to afford diphenylurea has been investigated. The reaction is best performed with equimolar amounts of the two reagents. Use of higher concentrations of either aniline or nitrobenzene or an increase in temperature in the range 120-170 °C leads to the formation of higher amounts of azo- and azoxybenzene. The latter were found to contain exclusively the aryl moiety deriving from nitrobenzene, with no inclusion of that derived from aniline. The addition of a small amount of diphenylphosphinic acid doubles the conversion and improves the selectivity in diphenylurea, but the effect is attenuated for larger amounts of acid. Small amounts of chloride, of the order of 10-30 mol% with respect to palladium, improve both rate and selectivity, but only inhibiting effects are detected when chloride is added to the reaction mixture for the carbonylation of 2,4-dinitrotoluene to dimethyl 2,4-toluenedicarbamate. The data obtained and that previously reported in the literature has been analyzed in the context of a unifying mechanism and an explanation for some apparent contradictions has been given.     

Synthesis of micelle templated silico-aluminas with different alumina contents

The computer aided analysis of the EPR spectra of radical surfactant probes inserted in cetyltrimethylammonium bromide micelles provided information on the kinetics of formation of micelle templated silico-aluminas (MTSA) at 343 K, obtained by means of silica and alumina alkaline solutions at different Si/Al ratios (from infinity to 4). Mainly two spectral components were analyzed and relatively quantified in the EPR spectra: (1) the micellar component, due to probes inserted in the surfactant aggregates, whose mobility decreases over the synthesis time, thus reporting on the progressive modification of the micelle structure and the solid condensation; (2) the interacting component, mainly arising from the electrostatic interactions between the surfactant heads and the charged surface sites. This last component increases its relative intensity over the synthesis time, informing about condensation and structuration of the silico-alumina at the micelle surface. X-ray diffraction (XRD), nitrogen sorption isotherms at 77 K, thermogravimetric analysis, TEM and chemical analysis were performed to characterize both as-synthesized and calcined MTSA materials. Nitrogen sorption isotherms allowed us to evaluate the pore diameter, the specific surface area and the pore volume. At Si/Al<15 a decrease in pore volume and specific surface area was interpreted as due to the contemporaneous presence of a hexagonal MTSA and an amorphous material, which was ascertained by means of XRD as the only present at Si/Al=4. The amorphous structure at Si/Al<15 used Na+ as contraions, whereas the surfactants are no more needed to neutralize the negatively charged groups at the solid surface. The hypothesis of a "break" at Si/Al=15 was supported by EPR: the interactions between the surfactant probe heads and the negatively charged surface groups are drastically reduced at Si/Al<15. On the contrary, at Si/Al>15, increasing amounts of alumina slow the kinetics of the synthesis but enhance electrostatic interactions between the surfactant heads and the negatively charged surface groups. Dilution of the synthesis mixture decreased the extent of the interactions, due to partial protonation of the silanol groups, and slowed the synthesis process.     

C2C12 myoblast sensitivity to different apoptotic chemical triggers

Apoptosis is a form of cell death crucial for normal development and tissue homeostasis. Its typical features include chromatin changes, nuclear breakdown, plasma membrane blebbing and splitting of cellular content into apoptotic bodies, that progressively undergo phagocytosis.Apoptosis is considered essential for skeletal muscle development, where defective cells are deleted during differentiation. In addition, it plays a relevant role in several muscle myopathies, as well as in denervation and disuse.The aim of this study was to evaluate muscle cell sensitivity to different apoptotic triggers, acting through different mechanisms of action. Chemical agents, active against distinct intracellular targets, such as mitochondrial respiratory chain and DNA, have been chosen to better highlight cell death mechanisms. To induce apoptosis, C2C12 myoblasts have been exposed to H₂O₂, staurosporine, cisplatin and etoposide, at different doses and incubation times, and they have been analysed by flow cytometry, scanning and transmission electron microscopy.Flow cytometry analysis revealed a certain subdiploid peak after all treatments. The best apoptotic effect was observable, as confirmed at reverted microscope, at minimum doses and after the major exposure time.At ultrastructural level programmed cell death has been observed. Characteristic chromatin condensation and margination, as well as apoptotic bodies, frequently appeared, even if in the presence of secondary necrosis; surface blebs were also observed during scanning microscopic observation.In particular, exposure to H₂O₂ or staurosporine showed the largest number of myoblasts in late apoptotic stages and in secondary necrosis. Cisplatin treatments revealed few early apoptotic cells. The analysis of etoposide-induced apoptosis was in agreement with data obtained from flow cytometry, indicating a significant increase of apoptotic cell number.These results suggest that all conditions are able to induce apoptosis in C2C12 myoblasts, which occurs, considering trigger mechanisms of action, mostly following the mitochondrial pathway, if not excluding that due to DNA damage. Therefore, mitochondria permeability alteration is an important step in skeletal muscle programmed cell death. This last conclusion seems to have a significant relevance in understanding the mechanisms involved in muscle disorders, denervation and chronic muscle disuse, conditions frequently characterized by a decline in mitochondrial content and by an increase of mitochondrial apoptosis susceptibility.     

Regularity of line configurations

We show that in arithmetically-Gorenstein line arrangements with only planar singularities, each line intersects the same number of other lines. This number has an algebraic interpretation: it is the Castelnuovo–Mumford regularity of the coordinate ring of the arrangement.We also prove that every $(d?1)$-dimensional simplicial complex whose 0-th and 1-st homologies are trivial is the nerve complex of a suitable d-dimensional standard graded algebra of depth ≥3. This provides the converse of a recent result by Katzman, Lyubeznik and Zhang.     

Mathematics learning and tools from theoretical, historical and practical points of view: the productive notion of mathematics laboratories

In our research work, we have looked at the way in which artefacts become, for teachers as well as for students, instruments of their mathematical activity. The issues related to the use of tools and technologies in mathematical education are now widely considered. A look to history highlights the different ways in which the same questions have been studied at different times and in different places. This suggests that the contribution of artefacts to mathematics learning should be considered in terms of various contexts. Our “visits” to these contexts will be guided by the coordination of two main theoretical frameworks, the instrumental approach and the semiotic mediation approach from the perspective of mathematics laboratory. This journey through history and schooling represents a good occasion to address some questions: Are there “good” contexts in which to develop mathematical instruments? Are there “good” teaching practices which assist students’ instrumental geneses and construct mathematical meanings? How is it possible to promote such teaching practices? Some study cases are discussed.