Binding affinity of fluorochromes and fluorescent proteins to Taxol™ crystals

The ability of Taxol (paclitaxel) to bind and stabilize microtubules is the basis for its use as an anti-mitotic drug as well as an additive for in vivo and in vitro studies of microtubules. The low solubility of Taxol in aqueous solutions, however, facilitates the formation of Taxol crystals that can be decorated with fluorescent tubulin. In cells treated with Taxol, these decorated Taxol crystals may be mistaken for self-assembled tubulin asters when observed with a fluorescent microscope. We confirmed via fluorescent and differential interference contrast microscopy that Taxol crystals can be decorated not only with fluorescent tubulin but also with other fluorescent proteins and fluorochromes without perturbing their morphology. We used theoretical calculations to further investigate Taxol-fluorescent agent interactions. Using computational docking studies we identified a new, potential Taxol binding site within the tubulin dimmer, allowing the interaction between crystalline Taxol and tubulin. Our calculations, however, show that fluorescent tubulin binding to Taxol crystals is more favorable via the fluorochromes covalently linked to the tubulin dimmers, rather than via the new Taxol-binding site, what is in accordance with our experimental data.     

Synthesis and characterization of a new family of photoactive liquid crystalline polyesters based on α‐methylstilbene

A liquid crystal polyester and two liquid crystal copolyesters containing α‐methylstilbene moieties and aliphatic or aromatic spacers in the backbone were synthesized in good yields, with the aim of using them for photosensitive microcapsule preparation. The synthesized polymers were fully characterized with respect to thermal stability, type of mesophase, molecular weight and E–Z photoisomerization. Combination of monomers with different structures allowed adjustment of the polymer characteristics such as degree of crystallinity and glass transition temperature, as verified using X‐ray diffraction, polarized optical microscopy and differential scanning calorimetry. Quantitative ¹H NMR and UV‐visible experiments were performed in order to investigate E–Z photoisomerization after photoirradiation at 364 nm. Finally, a membrane based on one of these polymers was prepared and it was found that its wettability increased on photoirradiation. © 2013 Society of Chemical Industry     

Structural and electrical properties of Cr-doped SrTiO3 porous materials

Series of single-phase materials with assumed formula SrTi_1?xCr_xO₃ (where x = 0, 1, 4, 6 mol.%) were obtained by sol-gel method. The structure and microstructure of materials were characterised by X-ray diffraction and scanning electron microscopy methods. Moreover, the study of electrical properties and evaluation of chemical stability in CO₂/H₂O atmosphere was performed by electrochemical impedance spectroscopy and thermogravimery methods, respectively. The possibility of participation of Cr-doped strontium titanate in oxidation–reduction processes was analysed by temperature-programed reduction (TPR) and temperature-programed oxidation (TPOx) measurements. The changes of lattice parameters together with XPS analysis, the Seebeck coefficient measurements results and TPR profiles obtained for SrTi_1?xCr_xO₃ materials prove the presence of chromium on +3 and +6 oxidation stages. Thus, chromium can be treated as both acceptor- and donor-type dopant in the SrTiO₃ structure. The Cr³⁺/Cr⁶⁺ ratio strongly affects the electrical properties, as the change of conduction mechanism was observed. The results of performed stability test clearly indicate that incorporation of chromium into SrTiO₃ structure results with decrease of chemical stability in CO₂ atmosphere.     

Synthesis and Characterization of Low Viscosity Dimethacrylic Resin Based on Isosorbide

In this article, synthesis and properties of novel dimethacrylic resin (ISETDMA) based on human friendly, biobased isosorbide was described. Its potential as a possible diluting monomer for medical applications, mainly dental restorative systems was assessed. The resin was obtained in two‐step synthesis including ethoxylation of isosorbide and subsequent methacrylation with methacryloyl chloride. ¹HNMR, FTIR, and electrospray ionization mass spectroscopy (ESI‐MS) techniques were used to identify products. ISETDMA as well as composition with 2,2‐bis[4‐(2‐hydroxy‐3‐methacryloyloxypropoxy)phenyl]propane were polymerized using UV initiator IRGACURE 651. Double bond conversion, polymerization shrinkage, water sorption, and sol fraction of resulting polymers were determined. Selected mechanical (flexural strength and modulus, Brinell hardness) and thermomechanical (dynamic mechanical analysis) properties were also investigated. Triethylene glycol dimethacrylate and 2,2‐bis(4‐(2‐methacryloxyethyl‐1‐oxy)phenyl)propane based homopolymers and copolymers were prepared as reference for comparison of particular properties. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 130: 2514–2522, 2013     

Graphoepitaxy of High‐Quality GaN Layers on Graphene/6H–SiC

The implementation of graphene layers in gallium nitride (GaN) heterostructure growth can solve self‐heating problems in nitride‐based high‐power electronic and light‐emitting optoelectronic devices. In the present study, high‐quality GaN layers are grown on patterned graphene layers and 6H–SiC by metalorganic chemical vapor deposition. A periodic pattern of graphene layers is fabricated on 6H–SiC by using polymethyl methacrylate deposition and electron beam lithography, followed by etching using an Ar/O₂ gas atmosphere. Prior to GaN growth, an AlN buffer layer and an Al_0.2Ga_0.8N transition layer are deposited. The atomic structures of the interfaces between the 6H–SiC and graphene, as well as between the graphene and AlN, are studied using scanning transmission electron microscopy. Phase separation of the Al_0.2Ga_0.8N transition layer into an AlN and GaN superlattice is observed. Above the continuous graphene layers, polycrystalline defective GaN is rapidly overgrown by better quality single‐crystalline GaN from the etched regions. The lateral overgrowth of GaN results in the presence of a low density of dislocations (≈10⁹ cm⁻²) and inversion domains and the formation of a smooth GaN surface.     

Predicted Ligands for the Human Urotensin‐II G Protein‐Coupled Receptor with Some Experimental Validation

Human Urotensin‐II (U‐II) is the most potent mammalian vasoconstrictor known. 1 Thus, a U‐II antagonist would be of therapeutic value in a number of cardiovascular disorders. 2 Here, we describe our work on the prediction of the structure of the human U‐II receptor (hUT₂R) using GEnSeMBLE (GPCR Ensemble of Structures in Membrane BiLayer Environment) complete sampling Monte Carlo method. With the validation of our predicted structures, we designed a series of new potential antagonists predicted to bind more strongly than known ligands. Next, we carried out R‐group screening to suggest a new ligand predicted to bind with 7 kcal mol^?1 better energy than 1‐{2‐[4‐(2‐bromobenzyl)‐4‐hydroxypiperidin‐1‐yl]ethyl}‐3‐(thieno[3,2‐b]pyridin‐7‐yl)urea, the designed antagonist predicted to have the highest affinity for the receptor. Some of these predictions were tested experimentally, validating the computational results. Using the pharmacophore generated from the predicted structure for hUT₂R bound to ACT‐058362, we carried out virtual screening based on this binding site. The most potent hit compounds identified contained 2‐(phenoxymethyl)‐1,3,4‐thiadiazole core, with the best derivative exhibiting an IC₅₀ value of 0.581 μM against hUT₂R when tested in vitro. Our efforts identified a new scaffold as a potential new lead structure for the development of novel hUT₂R antagonists, and the computational methods used could find more general applicability to other GPCRs.     

Polymeric Microspheres/Cells/Extracellular Matrix Constructs Produced by Auto-Assembly for Bone Modular Tissue Engineering

Modular tissue engineering (MTE) is a novel “bottom-up” approach to create engineered biological tissues from microscale repeating units. Our aim was to obtain microtissue constructs, based on polymer microspheres (MSs) populated with cells, which can be further assembled into larger tissue blocks and used in bone MTE. Poly(L-lactide-co-glycolide) MS of 165 ± 47 µm in diameter were produced by oil-in-water emulsification and treated with 0.1 M NaOH. To improve cell adhesion, MSs were coated with poly-L-lysine (PLL) or human recombinant collagen type I (COL). The presence of oxygenated functionalities and PLL/COL coating on MS was confirmed by X-ray photoelectron spectroscopy (XPS). To assess the influence of medium composition on adhesion, proliferation, and osteogenic differentiation, preosteoblast MC3T3-E1 cells were cultured on MS in minimal essential medium (MEM) and osteogenic differentiation medium (OSG). Moreover, to assess the potential osteoblast–osteoclast cross-talk phenomenon and the influence of signaling molecules released by osteoclasts on osteoblast cell culture, a medium obtained from osteoclast culture (OSC) was also used. To impel the cells to adhere and grow on the MS, anti-adhesive cell culture plates were utilized. The results show that MS coated with PLL and COL significantly favor the adhesion and growth of MC3T3-E1 cells on days 1 and 7, respectively, in all experimental conditions tested. On day 7, three-dimensional MS/cell/extracellular matrix constructs were created owing to auto-assembly. The cells grown in such constructs exhibited high activity of early osteogenic differentiation marker, namely, alkaline phosphatase. Superior cell growth on PLL- and COL-coated MS on day 14 was observed in the OSG medium. Interestingly, deposition of extracellular matrix and its mineralization was particularly enhanced on COL-coated MS in OSG medium on day 14. In our study, we developed a method of spontaneous formation of organoid-like MS-based cell/ECM constructs with a few millimeters in size. Such constructs may be regarded as building blocks in bone MTE.     

Immunonutritional Protease Inhibitors from T. durum and A. sativa Display Metabolic Similarities When Assayed on Human Macrophage-like Cells

This study evaluated the immunonutritional effects caused by protease inhibitors from Avena sativa and Triticum durum to human macrophage-like cells. Macrophages were exposed (3 h) to extracts obtained from flours, and mitochondrial-associated oxygen consumption rates and inflammatory, metabolic, and proteome adaptations were quantified. Mass spectrometry ‘m/z’ signals of the extracts obtained from T. durum and A. sativa revealed molecular weights of 18–35 kDa and 16–22 kDa, respectively, for the compounds present at highest concentrations. Extracts from T. durum exhibited lower susceptibility to degradation by gastrointestinal enzymes than those from A. sativa: 9.5% vs 20.2%. Despite their different botanical origin, both extracts increased TLR4 expression. Metabolic protein levels were indicative of a decreased glycolytic to lactate flux in cell cultures upon stimulation with A. sativa extracts, which improved mitochondrial respiration in relation to those from T. durum. Principal components analysis confirmed relative similarities between immune–metabolic events triggered by immunonutritional ingredients in T. durum and A. sativa. Collectively, immunonutritional effects help to interpret the differences between both crops, worsening or improving, macrophage immune reactivity (tolerogenicity), and better control of inflammatory processes.