High-Fat Diet Impairs Mouse Median Eminence: A Study by Transmission and Scanning Electron Microscopy Coupled with Raman Spectroscopy

Hypothalamic dysfunction is an initial event following diet-induced obesity, primarily involving areas regulating energy balance such as arcuate nucleus (Arc) and median eminence (ME). To gain insights into the early hypothalamic diet-induced alterations, adult CD1 mice fed a high-fat diet (HFD) for 6 weeks were studied and compared with normo-fed controls. Transmission and scanning electron microscopy and histological staining were employed for morphological studies of the ME, while Raman spectroscopy was applied for the biochemical analysis of the Arc-ME complex. In HFD mice, ME β2-tanycytes, glial cells dedicated to blood-liquor crosstalk, exhibited remarkable ultrastructural anomalies, including altered alignment, reduced junctions, degenerating organelles, and higher content of lipid droplets, lysosomes, and autophagosomes. Degenerating tanycytes also displayed an electron transparent cytoplasm filled with numerous vesicles, and they were surrounded by dilated extracellular spaces extending up to the subependymal layer. Consistently, Raman spectroscopy analysis of the Arc-ME complex revealed higher glycogen, collagen, and lipid bands in HFD mice compared with controls, and there was also a higher band corresponding to the cyanide group in the former compared to the last. Collectively, these data show that ME β₂-tanycytes exhibit early structural and chemical alterations due to HFD and reveal for the first-time hypothalamic cyanide presence following high dietary lipids consumption, which is a novel aspect with potential implications in the field of obesity.     

Detection of changes in the cellular composition of Salmonella enterica serovar Typhimurium in the presence of antimicrobial compound(s) of Lactobacillus strains using Fourier transform infrared spectroscopy

It was previously established that Lactobacillus fermentum ACA-DC 179, Lactobacillus plantarum ACA-DC 287 and Lactobacillus plantarum ACA-DC 2350 exhibit antimicrobial activity against Salmonella enterica serovar Typhimurium. In order to further investigate the killing effect of these microorganisms against Salmonella cells, we employed Fourier transform infrared spectroscopy (FT-IR). Salmonella cells were incubated with different concentrated lactobacilli supernatants and their FT-IR spectra were recorded. The second derivative transformation of the original spectra revealed changes in spectral regions corresponding to absorptions of major cellular constituents (e.g. cell wall, cell membrane, and proteins of the cell) among the Salmonella cells treated with the supernatants and those treated with the control samples. Principal component analysis of the second derivative transformed spectra showed that the yet unidentified antimicrobial compound(s) produced by the lactobacilli tested clearly interfered with the fatty acids of the cell membrane, as well as the polysaccharides of the cell wall in Salmonella cells, pointing towards a dual killing mode. Our study shed light for the first time in the anti-Salmonella activity of the particular Lactobacillus strains.     

Photooxidative degradation and conformational changes in syndiotactic polypropylene/clay nanocomposites

Syndiotactic polypropylene nanocomposites based on layered silicates in various proportions were subjected to prolonged (246 h) ultraviolet (UV)-c irradiation. Fourier transform infrared (FT-IR) spectroscopy was used in order to investigate the molecular alterations of the polymeric matrix during the UV exposure relative to the concentration of nanoclay. It was found that a significant increase of the helical conformation upon the irradiation took place as a result of scissions of the polymeric chains. In addition, a simultaneous increase in the crystallinity was verified by X-ray diffraction (XRD) measurements. Furthermore, a variety of photooxidation products were detected, among them carboxylic acids, ketones, gamma-lactones, and esters. We report in this paper the impact of the clay on the degradation mechanism of syndiotactic polypropylene mostly by the production of additional free radicals. Therefore, the relative intensities of the produced photooxidative species are affected drastically as a function of the concentration of the layered silicate present. Finally, an interaction of the carbonyl groups formed in the polymer upon irradiation with the polar groups of the layered silicates was confirmed both from infrared and UV-visible spectroscopic studies.     

A Non-Hazardous Deparaffinization Protocol Enables Quantitative Proteomics of Core Needle Biopsy-Sized Formalin-Fixed and Paraffin-Embedded (FFPE) Tissue Specimens

Most human tumor tissues that are obtained for pathology and diagnostic purposes are formalin-fixed and paraffin-embedded (FFPE). To perform quantitative proteomics of FFPE samples, paraffin has to be removed and formalin-induced crosslinks have to be reversed prior to proteolytic digestion. A central component of almost all deparaffinization protocols is xylene, a toxic and highly flammable solvent that has been reported to negatively affect protein extraction and quantitative proteome analysis. Here, we present a ‘green’ xylene-free protocol for accelerated sample preparation of FFPE tissues based on paraffin-removal with hot water. Combined with tissue homogenization using disposable micropestles and a modified protein aggregation capture (PAC) digestion protocol, our workflow enables streamlined and reproducible quantitative proteomic profiling of FFPE tissue. Label-free quantitation of FFPE cores from human ductal breast carcinoma in situ (DCIS) xenografts with a volume of only 0.79 mm³ showed a high correlation between replicates (r² = 0.992) with a median %CV of 16.9%. Importantly, this small volume is already compatible with tissue micro array (TMA) cores and core needle biopsies, while our results and its ease-of-use indicate that further downsizing is feasible. Finally, our FFPE workflow does not require costly equipment and can be established in every standard clinical laboratory.     

The Interplay between the Theories of Mode Coupling and of Percolation Transition in Attractive Colloidal Systems

In the recent years a considerable effort has been devoted to foster the understanding of the basic mechanisms underlying the dynamical arrest that is involved in glass forming in supercooled liquids and in the sol-gel transition. The elucidation of the nature of such processes represents one of the most challenging unsolved problems in the field of material science. In this context, two important theories have contributed significantly to the interpretation of these phenomena: the Mode-Coupling theory (MCT) and the Percolation theory (PT). These theories are rooted on the two pillars of statistical physics, universality and scale laws, and their original formulations have been subsequently modified to account for the fundamental concepts of Energy Landscape (EL) and of the universality of the fragile to strong dynamical crossover (FSC). In this review, we discuss experimental and theoretical results, including Molecular Dynamics (MD) simulations, reported in the literature for colloidal and polymer systems displaying both glass and sol-gel transitions. Special focus is dedicated to the analysis of the interferences between these transitions and on the possible interplay between MCT and PT. By reviewing recent theoretical developments, we show that such interplay between sol-gel and glass transitions may be interpreted in terms of the extended F13 MCT model that describes these processes based on the presence of a glass-glass transition line terminating in an A3 cusp-like singularity (near which the logarithmic decay of the density correlator is observed). This transition line originates from the presence of two different amorphous structures, one generated by the inter-particle attraction and the other by the pure repulsion characteristic of hard spheres. We show here, combining literature results with some new results, that such a situation can be generated, and therefore experimentally studied, by considering colloidal-like particles interacting via a hard core plus an attractive square well potential. In the final part of this review, scaling laws associated both to MCT and PT are applied to describe, by means of these two theories, the specific viscoelastic properties of some systems.     

Using computer supported collaborative learning strategies for helping students acquire self-regulated problem-solving skills in mathematics

The main objective of this paper is to investigate the effectiveness of a proposed computer-based instructional method in Primary Education for self-regulated problem solving. The proposed instructional method is based on Sternberg’s model of problem solving within an authentic context. It consists of three main phases: observation, collaboration and semi-structured guidance. The ultimate learning objective is to augment the autonomous problem-solving skills of primary school children. In our study the Synergo tool was used, which is a synchronous computer supported collaborative learning tool, as well as the Moodle learning management system. The context which frames the method is authentic, very close to a students’ realistic learning situation. The findings of this study advocate that students can increase their problem-solving skills in a relatively short period of time. At the same time, they can improve their approach to the solution of a given mathematical problem, performing significant signs of autonomy.     

Polymeric Nanoparticles as a Self-Adjuvanting Peptide Vaccine Delivery System: The Role of Shape

Antigens incorporated in subunit vaccines are typically poorly immunogenic, so a strong immunostimulant (adjuvant) and/or delivery system is required to boost immunogenicity. In this work, the various functional polymer nanostructures, that is, rods, worms, spheres, and tadpoles are used to develop potent peptide antigen delivery systems. The antigen PADRE-J8 (PJ8), derived from Group A Streptococcus (GAS) M-protein, is either physically mixed or chemically conjugated to polymeric nanoparticles of different shapes. The physical mixture of polymeric nanoparticles and antigen is more effective in inducing antibody production than their chemical conjugates. Moreover, rod-shaped polymeric nanoparticles in physical mixture with PJ8 elicited higher and more opsonic antibody titers than powerful complete Freund's adjuvant (CFA)-adjuvanted antigen. Herein, for the first time it is demonstrated that a) the block copolymer, in nanoparticle form, can act as an immune adjuvant, b) nanoparticle shape plays a crucial role in their immunogenicity, and c) antigen conjugation is not required, nor is antigen encapsulation or absorption.