Quantification of Dendritic Spines Remodeling under Physiological Stimuli and in Pathological Conditions

Numerous brain diseases are associated with abnormalities in morphology and density of dendritic spines, small membranous protrusions whose structural geometry correlates with the strength of synaptic connections. Thus, the quantitative analysis of dendritic spines remodeling in microscopic images is one of the key elements towards understanding mechanisms of structural neuronal plasticity and bases of brain pathology. In the following article, we review experimental approaches designed to assess quantitative features of dendritic spines under physiological stimuli and in pathological conditions. We compare various methodological pipelines of biological models, sample preparation, data analysis, image acquisition, sample size, and statistical analysis. The methodology and results of relevant experiments are systematically summarized in a tabular form. In particular, we focus on quantitative data regarding the number of animals, cells, dendritic spines, types of studied parameters, size of observed changes, and their statistical significance.     

Radiation- and Photo-Induced Oxidation Pathways of Methionine in Model Peptide Backbone under Anoxic Conditions

Within the reactive oxygen species (ROS) generated by cellular metabolisms, hydroxyl radicals (HO^•) play an important role, being the most aggressive towards biomolecules. The reactions of HO^• with methionine residues (Met) in peptides and proteins have been intensively studied, but some fundamental aspects remain unsolved. In the present study we examined the biomimetic model made of Ac-Met-OMe, as the simplest model peptide backbone, and of HO^• generated by ionizing radiation in aqueous solutions under anoxic conditions. We performed the identification and quantification of transient species by pulse radiolysis and of final products by LC-MS and high-resolution MS/MS after γ-radiolysis. By parallel photochemical experiments, using 3-carboxybenzophenone (CB) triplet with the model peptide, we compared the outcomes in terms of short-lived intermediates and stable product identification. The result is a detailed mechanistic scheme of Met oxidation by HO^•, and by CB triplets allowed for assigning transient species to the pathways of products formation.     

Role of Polyinosinic:Polycytidylic Acid-Induced Maternal Immune Activation and Subsequent Immune Challenge in the Behaviour and Microglial Cell Trajectory in Adult Offspring: A Study of the Neurodevelopmental Model of Schizophrenia

Multiple lines of evidence support the pathogenic role of maternal immune activation (MIA) in the occurrence of the schizophrenia-like disturbances in offspring. While in the brain the homeostatic role of neuron-microglia protein systems is well documented, the participation of the CX3CL1-CX3CR1 and CD200-CD200R dyads in the adverse impact of MIA often goes under-recognized. Therefore, in the present study, we examined the effect of MIA induced by polyinosinic:polycytidylic acid (Poly I:C) on the CX3CL1-CX3CR1 and CD200-CD200R axes, microglial trajectory (MhcII, Cd40, iNos, Il-1β, Tnf-α, Il-6, Arg1, Igf-1, Tgf-β and Il-4), and schizophrenia-like behaviour in adult male offspring of Sprague-Dawley rats. Additionally, according to the “two-hit” hypothesis of schizophrenia, we evaluated the influence of acute challenge with Poly I:C in adult prenatally MIA-exposed animals on the above parameters. In the present study, MIA evoked by Poly I:C injection in the late period of gestation led to the appearance of schizophrenia-like disturbances in adult offspring. Our results revealed the deficits manifested as a diminished number of aggressive interactions, presence of depressive-like episodes, and increase of exploratory activity, as well as a dichotomy in the sensorimotor gating in the prepulse inhibition (PPI) test expressed as two behavioural phenotypes (MIA_PPI-low and MIA_PPI-high). Furthermore, in the offspring rats subjected to a prenatal challenge (i.e., MIA) we noticed the lack of modulation of behavioural changes after the additional acute immune stimulus (Poly I:C) in adulthood. The important finding reported in this article is that MIA affects the expression and levels of the neuron-microglia proteins in the frontal cortex and hippocampus of adult offspring. We found that the changes in the CX3CL1-CX3CR1 axis could affect microglial trajectory, including decreased hippocampal mRNA level of MhcII and elevated cortical expression of Igf-1 in the MIA_PPI-high animals and/or could cause the up-regulation of an inflammatory response (Il-6, Tnf-α, iNos) after the “second hit” in both examined brain regions and, at least in part, might differentiate behavioural disturbances in adult offspring. Consequently, the future effort to identify the biological background of these interactions in the Poly I:C-induced MIA model in Sprague-Dawley rats is desirable to unequivocally clarify this issue.     

Chloroquine-Induced Accumulation of Autophagosomes and Lipids in the Endothelium

Chloroquine (CQ) is an antimalarial drug known to inhibit autophagy flux by impairing autophagosome–lysosome fusion. We hypothesized that autophagy flux altered by CQ has a considerable influence on the lipid composition of endothelial cells. Thus, we investigated endothelial responses induced by CQ on human microvascular endothelial cells (HMEC-1). HMEC-1 cells after CQ exposure were measured using a combined methodology based on label-free Raman and fluorescence imaging. Raman spectroscopy was applied to characterize subtle chemical changes in lipid contents and their distribution in the cells, while the fluorescence staining (LipidTox, LysoTracker and LC3) was used as a reference method. The results showed that CQ was not toxic to endothelial cells and did not result in the endothelial inflammation at concentrations of 1–30 µM. Notwithstanding, it yielded an increased intensity of LipidTox, LysoTracker, and LC3 staining, suggesting changes in the content of neutral lipids, lysosomotropism, and autophagy inhibition, respectively. The CQ-induced endothelial response was associated with lipid accumulation and was characterized by Raman spectroscopy. CQ-induced autophagosome accumulation in the endothelium is featured by a pronounced alteration in the lipid profile, but not in the endothelial inflammation. Raman-based assessment of CQ-induced biochemical changes offers a better understanding of the autophagy mechanism in the endothelial cells.     

New 1,3,4-Oxadiazole Derivatives of Pyridothiazine-1,1-Dioxide with Anti-Inflammatory Activity

Numerous studies have confirmed the coexistence of oxidative stress and inflammatory processes. Long-term inflammation and oxidative stress may significantly affect the initiation of the neoplastic transformation process. Here, we describe the synthesis of a new series of Mannich base-type hybrid compounds containing an arylpiperazine residue, 1,3,4-oxadiazole ring, and pyridothiazine-1,1-dioxide core. The synthesis was carried out with the hope that the hybridization of different pharmacophoric molecules would result in a synergistic effect on their anti-inflammatory activity, especially the ability to inhibit cyclooxygenase. The obtained compounds were investigated in terms of their potencies to inhibit cyclooxygenase COX-1 and COX-2 enzymes with the use of the colorimetric inhibitor screening assay. Their antioxidant and cytotoxic effect on normal human dermal fibroblasts (NHDF) was also studied. Strong COX-2 inhibitory activity was observed after the use of TG6 and, especially, TG4. The TG11 compound, as well as reference meloxicam, turned out to be a preferential COX-2 inhibitor. TG12 was, in turn, a non-selective COX inhibitor. A molecular docking study was performed to understand the binding interaction of compounds at the active site of cyclooxygenases.     

Bacteriophages vB_Sen-TO17 and vB_Sen-E22, Newly Isolated Viruses from Chicken Feces,Specific for Several Salmonella enterica Strains

Two newly discovered bacteriophages, isolated from chicken feces and infecting Salmonella enterica strains, are described in this report. These phages have been named vB_Sen-TO17 and vB_Sen-E22, and we present their molecular and functional characterization. Both studied viruses are able to infect several S. enterica strains and develop lytically, but their specific host ranges differ significantly. Electron microscopic analyses of virions have been performed, and full genome sequences were determined and characterized, along with molecular phylogenetic studies. Genomes of vB_Sen-TO17 (ds DNA of 41,658 bp) and vB_Sen-E22 (dsDNA of 108,987 bp) are devoid of homologs of any known or putative gene coding for toxins or any other proteins potentially deleterious for eukaryotic cells. Both phages adsorbed efficiently (>95% adsorbed virions) within 10 min at 42 °C (resembling chicken body temperature) on cells of most tested host strains. Kinetics of lytic development of vB_Sen-TO17 and vB_Sen-E22, determined in one-step growth experiments, indicated that development is complete within 30–40 min at 42 °C, whereas burst sizes vary from 9 to 79 progeny phages per cell for vB_Sen-TO17 and from 18 to 64 for vB_Sen-E22, depending on the host strain. Virions of both phages were relatively stable (from several percent to almost 100% survivability) under various conditions, including acidic and alkaline pH values (from 3 to 12), temperatures from −80 °C to 60 °C, 70% ethanol, chloroform, and 10% DMSO. These characteristics of vB_Sen-TO17 and vB_Sen-E22 indicate that these phages might be considered in further studies on phage therapy, particularly in attempts to eliminate S. enterica from chicken intestine.     

Effects of Synthesis Conditions on the Formation of Si-Substituted Alpha Tricalcium Phosphates

Powders of α-TCP containing various amounts of silicon were synthesized by two different methods: Wet chemical precipitation and solid-state synthesis. The obtained powders were then physico–chemically studied using different methods: Scanning and transmission electron microscopy (TEM and SEM), energy-dispersive X-ray spectroscopy (EDS), powder X-ray diffractometry (PXRD), infrared and Raman spectroscopies (FT-IR and R), and solid-state nuclear magnetic resonance (ssNMR). The study showed that the method of synthesis affects the morphology of the obtained particles, the homogeneity of crystalline phase and the efficiency of Si substitution. Solid-state synthesis leads to particles with a low tendency to agglomerate compared to the precipitation method. However, the powders obtained by the solid-state method are less homogeneous and contain a significant amount of other crystalline phase, silicocarnotite (up to 7.33%). Moreover, the microcrystals from this method are more disordered. This might be caused by more efficient substitution of silicate ions: The silicon content of the samples obtained by the solid-state method is almost equal to the nominal values.     

Determination of Acoustical Parameters of Aqueous Solution of Kolliphors Binary Mixtures Using Density,Speed of Sound,Viscosity, and Surface Tension Measurements

Measurements of density, speed of sound, and surface tension as well as viscosity of aqueous Kolliphor® ELP (ELP) and Kolliphor® RH 40 (RH40) solutions as well as binary mixtures at different surfactant mole fractions were made at 293 K to investigate their aggregation behavior. The free volume, internal pressure, and molar cohesive energy were calculated and compared to the specific acoustic impedance and intermolecular free length to obtain qualitative information about the character of interactions between the surfactant molecules in the mixture through the water phase.     

Improved photon management in a photoelectrochemical cell with Nd-modified TiO2 thin film photoanode

Photon management involving particularly an up-conversion process is proposed as a relatively novel strategy for improving the efficiency of hydrogen generation in photoelectrochemical cells (PEC) with wide-band gap photoanodes. Optically active photoanode has been constructed by electrodeposition of titanium dioxide nanopowders containing Nd³⁺ ions, synthesized via a sol-gel method, onto ITO/TiO₂(thin film) substrates. Thin films of TiO₂ have been deposited by means of RF magnetron sputtering in an ultra-high-vacuum system. X-ray diffraction, scanning electron microscopy, UV-VIS-NIR spectrophotometry, and photoluminescence have been applied to assess the properties of photoanodes. In experiments involving photon-assisted water splitting, an external up-converter containing Yb³⁺/Er³⁺ rare-earth ions has been used. Photocurrent as a function of voltage (V_B) under illumination with white light is relatively high (280 μA at V_B = 0 V) for pure TiO₂ thin films and it is not affected by the electrodeposition of TiO₂:Nd³⁺ powders. NIR-driven up-conversion with laser excitation at λ = 980 nm has been found responsible for a 13-fold increase in photocurrent at V_B = 0 V in the modified PEC configuration.