Elastomeric Protein Bioactive Eutectogels for Topical Drug Delivery

Therapeutic deep eutectic solvents (THEDES) are an emerging family of eutectic mixtures gaining increasing interest in the biomedical space. The immobilization of THEDES into polymer networks allows bioactive eutectogels to expand their application scope to topical drug delivery. Herein, this work presents the first set of elastomeric eutectogels constructed by supporting a therapeutic eutectic system with skin permeation ability in a protein scaffold dynamically crosslinked by a natural polyphenol. In this ionic eutectic, gelatin undergoes gelation through an unexpected mechanism in striking contrast with classical hydrogels, which is herein thoroughly studied. Interestingly, the polyphenol controls the conformation of the protein structure, enabling tuning up the mechanical and viscoelastic behavior of the dynamic eutectogel networks from elastic to hyperelastic. The resultant protein eutectogels exhibit strain-hardening behavior, thermoreversible gel-to-sol transition, and excellent adhesive performance. Furthermore, these versatile materials retain the bioactivity of the liquid THEDES and favor skin occlusion, assisting the delivery of both hydrophilic and hydrophobic substances in ex vivo porcine skin in a time-dependent penetration process. These ultrastretchable eutectogels show new interplays between protein scaffolds and eutectic mixtures, paving the way for innovative therapeutic soft materials.     

Pd(0)-Catalyzed-Mediated Synthesis of Vitamin D Compounds

1α,25-Dihydroxyvitamin D₃ (1,25D₃), the hormonally active form of vitamin D₃, regulates mineral homeostasis, cell growth, cell differentiation-proliferation, apoptosis, and immune responses. 1,25D₃ activates more than 229 genes associated with several diseases, including arthritis, diabetes and cancer, suggesting its implications in a broader range of biological functions than originally thought. Despite the wide range of biological activities, the clinical applications of 1,25D₃ have been limited due to its collateral hypercalcemic effects. This problem has boosted intense synthetic activity in the vitamin D area in the last decades aimed at the development of highly active and non-calcemic analogs for treatment of hyperproliferative diseases. This review covers the most useful synthetic approaches to 1,25D₃ analogs containing the natural vitamin D triene system with emphasis on the Pd(0)-catalyzed transformations involved in the formation of the vitamin D triene system and A-ring synthons.     

A generalized global adaptive tracking control scheme for robot manipulators with bounded inputs

In this work, a generalized adaptive scheme for the global motion control of robot manipulators with constrained inputs is proposed. It gives rise to various families of bounded adaptive controllers defined through a general class of saturation functions. Compared with adaptive tracking control algorithms previously developed in a bounded input context, the proposed adaptive approach guarantees the motion control objective for any initial condition, avoiding discontinuities throughout the scheme, preventing the inputs to reach their natural saturation bounds, and permitting innovation on the saturating structure through its generalized form, giving a wide range of possibilities for performance improvement. Experimental results corroborate the efficiency of the proposed scheme. Copyright © 2014 John Wiley & Sons, Ltd.     

A new combined computational and NMR-spectroscopical strategy for the identification of additional conformational constraints of the bound ligand in an aprotic solvent

This study documents the feasibility of switching to an aprotic medium in sugar receptor research. The solvent change offers additional insights into mechanistic details of receptor--carbohydrate ligand interactions. If a receptor retained binding capacity in an aprotic medium, solvent-exchangeable protons of the ligand would not undergo transfer and could act as additional sensors, thus improving the level of reliability in conformational analysis. To probe this possibility, we first focused on hevein, the smallest lectin found in nature. The NMR-spectroscopic measurements verified complexation, albeit with progressively reduced affinity by more than 1.5 orders of magnitude, in mixtures of up to 50% dimethyl sulfoxide (DMSO). Since hevein lacks the compact beta-strand arrangement of other sugar receptors, such a structural motif may confer enhanced resistance to solvent exchange. Two settings of solid-phase activity assays proved this assumption for three types of alpha- and/or beta-galactoside-binding proteins, that is, a human immunoglobulin G (IgG) subfraction, the mistletoe lectin, and a member of the galectin family of animal lectins. Computer-assisted calculations and NMR experiments also revealed no conspicuous impact of the solvent on the conformational properties of the tested ligands. To define all possible nuclear Overhauser effect (NOE) contacts in a certain conformation and to predict involvement of exchangeable protons, we established a new screening protocol applicable during a given molecular dynamics (MD) trajectory and calculated population densities of distinct contacts. Experimentally, transferred NOE (tr-NOE) experiments with IgG molecules and the disaccharide Gal'alpha1-3Galbeta1-R in DMSO as solvent disclosed that such an additional crosspeak, that is, Gal'OH2--GalOH4, was even detectable for the bound ligand under conditions in which spin diffusion effects are suppressed. Further measurements with the plant lectin and galectins confirmed line broadening of ligand signals and gave access to characteristic crosspeaks in the aprotic solvent and its mixtures with water. Our combined biochemical, computational, and NMR-spectroscopical strategy is expected to contribute notably to the precise elucidation of the geometry of ligands bound to compactly folded sugar receptors and of the role of water molecules in protein--ligand (carbohydrate) recognition, with relevance to areas beyond the glycosciences.     

Differential item functioning in the Danish translation of the SF-36

Statistical analyses of Differential Item Functioning (DIF) can be used for rigorous translation evaluations. DIF techniques test whether each item functions in the same way, irrespective of the country, language, or culture of the respondents. For a given level of health, the score on any item should be independent of nationality. This requirement can be tested through contingency-table methods, which are efficient for analyzing all types of items. We investigated DIF in the Danish translation of the SF-36 Health Survey, using two general population samples (USA, n = 1,506; Denmark, n = 3,950). DIF was identified for 12 out of 35 items. These results agreed with independent ratings of translation quality, but the statistical techniques were more sensitive. When included in scales, the items exhibiting DIF had only a little impact on conclusions about cross-national differences in health in the general population. However, if used as single items, the DIF items could seriously bias results from cross-national comparisons. Also, the DIF items might have larger impact on cross-national comparison of groups with poorer health status. We conclude that analysis of DIF is useful for evaluating questionnaire translations.     

Proteomics identification of differentially expressed proteins in the muscle of dysferlin myopathy patients

The muscular dystrophies are a large and heterogeneous group of neuromuscular disorders that can be classified according to the mode of inheritance, the clinical phenotype and the molecular defect. To better understand the pathological mechanisms of dysferlin myopathy we compared the protein-expression pattern in the muscle biopsies of six patients with this disease with six patients with limb girdle muscular dystrophy 2A, five with facioscapulohumeral dystrophy and six normal control subjects. To investigate differences in the expression levels of skeletal muscle proteins we used 2-DE and MS. Western blot or immunohistochemistry confirmed relevant results. The study showed specific increase expression of proteins involved in fast-to-slow fiber type conversion (ankyrin repeat protein 2), type I predominance (phosphorylated forms of slow troponin T), sarcomere stabilization (actinin-associated LIM protein), protein ubiquitination (TRIM 72) and skeletal muscle differentiation (Rho-GDP-dissociation inhibitor ly-GDI) in dysferlin myopathy. As anticipated, we also found differential expression of proteins common to all the muscular dystrophies studied. This comparative proteomic analysis suggests that in dysferlin myopathy (i) the type I fiber predominance is an active process of fiber type conversion rather than a selective loss of type II fibers and (ii) the dysregulation of proteins involved in muscle differentiation further confirms the role of dysferlin in this process.     

Role of Methyl Salicylate on Oviposition Deterrence in Arabidopsis thaliana

Plants attacked by herbivores have evolved different strategies that fend off their enemies. Insect eggs deposited on leaves have been shown to inhibit further oviposition through visual or chemical cues. In some plant species, the volatile methyl salicylate (MeSA) repels gravid insects but whether it plays the same role in the model species Arabidopsis thaliana is currently unknown. Here we showed that Pieris brassicae butterflies laid fewer eggs on Arabidopsis plants that were next to a MeSA dispenser or on plants with constitutively high MeSA emission than on control plants. Surprisingly, the MeSA biosynthesis mutant bsmt1-1 treated with egg extract was still repellent to butterflies when compared to untreated bsmt1-1. Moreover, the expression of BSMT1 was not enhanced by egg extract treatment but was induced by herbivory. Altogether, these results provide evidence that the deterring activity of eggs on gravid butterflies is independent of MeSA emission in Arabidopsis, and that MeSA might rather serve as a deterrent in plants challenged by feeding larvae.     

Gossypol disrupts embryo development in heifers

Our objectives were to determine the effects of dietary free gossypol (FG) intake on plasma and uterine gossypol concentrations and embryo development and viability before and after culture with gossypol. Fifty postpubertal Holstein heifers weighing (±SD) 406 ± 34.5 kg at 11.5 mo of age were blocked by age and body weight (BW) and randomly assigned to 1 of 3 isocaloric and isonitrogenous diets differing in their FG content: control (0 mg of FG/kg of BW), moderate (17.8 mg of FG/kg of BW), and high (36.8 mg of FG/kg of BW). Heifers were fed the diets for 70 d before superovulation and embryo collection. Superovulated heifers were flushed on d 5 after induction of ovulation, and early morulae were either stained, to determine the number and proportion of live and dead cells, or randomly assigned to an in vitro culture for 96 h in media containing either 0 or 10 μg/mL of gossypol acetic acid. Plasma and uterine gossypol concentrations increased with increasing gossypol intake. The number of low-quality embryos-ova was greater for the high than for the moderate and control diets. Embryos collected from the high diet had the least number of cells because of fewer live cells, and were smaller in diameter. Greater dietary gossypol reduced blastocyst development and extended the time to reach the blastocyst stage. Similarly, gossypol concentration at 10 μg/mL compromised in vitro development and increased the proportion of degenerated embryos at 96 h in culture. These findings provide in vivo and in vitro evidence that intake of 36.8 mg of FG/kg of BW per d and gossypol concentrations >7 μg/mL in plasma, in uterine flush, or in vitro compromise early embryo development, which might explain some of the negative effects of gossypol on the fertility of dairy cows.     

Iron, zinc and copper content of foods commonly consumed in Mexico

Nutrient composition in foods is necessary for the determination of nutrient intake. Food composition tables used for dietary studies in Mexico do not have information of zinc, iron and copper; when present the values have been extrapolated from laboratory analysis carried out with foods in other countries and regions of the world. In this study zinc, iron, and copper content of 104 plant foods and 32 animal foods was determined. The procedure used was atomic absorption spectrophotometry for the minerals. Foods were grouped into cereals, vegetables, fruits, legumes and animal foods. Zinc content ranged from 0.018 mg/100 g for strawberry to 9.193 mg/100 g for beef. Iron content ranged from 0.113 mg/100 g for yogurt to 19.82 mg/100 g for a commercial cereal which had minerals added during processing. In some foods copper was not found and the highest content was 3.371 mg/100 g in beef liver. This study has provided information on zinc, iron and copper content of the most commonly consumed foods in Mexico.     

Improvements on enzymatic hydrolysis of human hair for illicit drug determination by gas chromatography/mass spectrometry

The use of ultrasound energy for accelerating the pronase E enzymatic hydrolysis of human hair for extracting illicit drugs has been novelty tested. The enzymatic extracts obtained after 30 min of sonication in an ultrasonic water bath were subjected to an optimized solid-phase extraction process, which involved a solution of 2.0% (v/v) acetic acid in methanol as eluting solution and concentration by N2 stream evaporation. A gas chromatography/mass spectrometry method was used to separate and determine cocaine, benzoylecgonine, codeine, morphine, and 6-monoacethylmorphine in 20 min. Variables affecting ultrasound-assisted pronase E hydrolysis such as hydrolysis temperature, hydrolysis time, enzyme concentration, catalyzer (1,4-dithiothreitol) concentration, ionic strength, pH, and ultrasound frequency were simultaneously evaluated by a Plackett-Burman design 2(8) PBD of resolution III. The most statistically significant variables were ionic strength and pH, which means that analyte extraction is mainly attributed to pronase E activity. The optimization or evaluation of all the factors has led to an accelerated pronase E hydrolysis of human hair, which can be completed in 30 min. Results have been found to be statistically similar to those obtained with conventional pronase E hydrolysis. The accelerated method was finally applied to several human hair samples from multidrug abusers.