Highly Conductive Fe-Doped (La,Sr)(Ga,Mg)O3-δ Solid-State Membranes for Electrochemical Application.

Membranes based on complex solid oxides with oxygen-ionic conductivity are widely used in high-temperature electrochemical devices such as fuel cells, electrolyzers, sensors, gas purifiers, etc. The performance of these devices depends on the oxygen-ionic conductivity value of the membrane. Highly conductive complex oxides with the overall composition of (La,Sr)(Ga,Mg)O3 have regained the attention of researchers in recent years due to the progress in the development of electrochemical devices with symmetrical electrodes. In this research, we studied how the introduction of iron cations into the gallium sublattice in (La,Sr)(Ga,Mg)O3 affects the fundamental properties of the oxides and the electrochemical performance of cells based on (La,Sr)(Ga,Fe,Mg)O3. It was found that the introduction of iron leads to an increase in the electrical conductivity and thermal expansion in an oxidizing atmosphere, while no such behavior was observed in a wet hydrogen atmosphere. The introduction of iron into a (La,Sr)(Ga,Mg)O3 electrolyte leads to an increase in the electrochemical activity of Sr2Fe1.5Mo0.5O6-δ electrodes in contact with the electrolyte. Fuel cell studies have shown that, in the case of a 550 µm-thick Fe-doped (La,Sr)(Ga,Mg)O3 supporting electrolyte (Fe content 10 mol.%) and symmetrical Sr2Fe1.5Mo0.5O6-δ electrodes, the cell exhibits a power density of more than 600 mW/cm2 at 800 °C.

Identification of New FG-Repeat Nucleoporins with Amyloid Properties.

Amyloids are fibrillar protein aggregates with a cross-ß structure. More than two hundred different proteins with amyloid or amyloid-like properties are already known. Functional amyloids with conservative amyloidogenic regions were found in different organisms. Protein aggregation appears to be beneficial for the organism in these cases. Therefore, this property might be conservative for orthologous proteins. The amyloid aggregates of the CPEB protein were suggested to play an important role in the long-term memory formation in Aplysia californica, Drosophila melanogaster, and Mus musculus. Moreover, the FXR1 protein demonstrates amyloid properties among the Vertebrates. A few nucleoporins (e.g., yeast Nup49, Nup100, Nup116, and human Nup153 and Nup58), are supposed or proved to form amyloid fibrils. In this study, we performed wide-scale bioinformatic analysis of nucleoporins with FG-repeats (phenylalanine-glycine repeats). We demonstrated that most of the barrier nucleoporins possess potential amyloidogenic properties. Furthermore, the aggregation-prone properties of several Nsp1 and Nup100 orthologs in bacteria and yeast cells were analyzed. Only two new nucleoporins, Drosophila melanogaster Nup98 and Schizosaccharomyces pombe Nup98, aggregated in different experiments. At the same time, Taeniopygia guttata Nup58 only formed amyloids in bacterial cells. These results rather contradict the hypothesis about the functional aggregation of nucleoporins.

Evaluation of the potential defensive strategy against Influenza A in cell line models.

Background: Influenza virus can cause both seasonal infections and unpredictable pandemics. Rapidly evolving avian H5N1 and  H7N9 viruses have a potential pandemic threat for humans. Since avian Influenza can be transmitted by domestic birds, serving as a key link between wild birds and humans, an effective measure to control the influenza transmission would be eradication of the infection in poultry. It is known that the virus penetrates into the cell through binding with the terminal oligosaccharides - sialic acids (SA) - on the cell surfaces. Removal of SA might be a potential antiviral strategy. An approach to developing chicken lines that are resistant to influenza viruses could be the creation of genetically modified birds. Thus it is necessary to select a gene that provides defense to influenza. Here we have expressed in cells a range of exogenous sialidases and estimated their activity and specificity towards SA residues. Methods: Several bacterial, viral and human sialidases were tested. We adopted bacterial sialidases from Salmonella and Actinomyces for expression on the cell surface by fusing catalytic domains with transmembrane domains. We also selected Influenza A/PuertoRico/8/34/H1N1 neuraminidase and human membrane sialidase ( hNeu3) genes. Lectin binding assay was used for estimation of a α (2,3)-sialylation level by fluorescent microscopy and FACS.   Results: We compared sialidases from bacteria, Influenza virus and human. Sialidases from Salmonella and Influenza A neuraminidase effectively cleaved α (2-3)-SA receptors. Viral neuraminidase demonstrated a higher activity. Sialidases from Actinomyces and hNeu3 did not show any activity against α (2-3) SA under physiological conditions. Conclusion: Our results demonstrated that sialidases with different specificity and activity can be selected as genes providing antiviral defence. Combining chosen sialidases with different activity together with tissue-specific promoters would provide an optimal level of desialylation. Tissue specific expression of the sialidases could protect domestic birds from infection.

Successful CRISPR/Cas9 mediated homologous recombination in a chicken cell line.

Background: CRISPR/Cas9 system is becoming the dominant genome editing tool in a variety of organisms. CRISPR/Cas9 mediated knock out has been demonstrated both in chicken cell lines and in chicken germ cells that served to generate genetically modified birds. However, there is limited data about CRISPR/Cas9 dependent homology directed repair (HDR) for avian, even in cell culture. Few attempts have been made with integrations in safe harbor loci of chicken genome that induces constitutive expression of the inserted gene. Gene expression under an endogenous promoter would be more valuable than under a constitutive exogenous promoter, as it allows the gene expression to be tissue-specific. Methods: Three gRNAs were chosen to target chicken 3'-untranslated region of GAPDH gene. Cas9-mediated activity in the targeted locus for the gRNAs in DF-1 cells was estimated by T7E1 assay. To edit the locus, the HDR cassette was added along with CRISPR/Cas9. The inserted sequence contained eGFP in frame with a GAPDH coding sequence via P2A and Neomycin resistance gene ( neoR) under cytomegalovirus promoter. Correct integration of the cassette was confirmed with fluorescent microscopy, PCR analysis and sequencing. Enrichment of modified cells was done by G418 selection. Efficiency of integration was assessed with fluorescence activated cell sorting (FACS). Results: We have established a CRISPR/Cas9 system to target an endogenous locus and precisely insert a gene under endogenous control. In our system, we used positive and negative selection to enrich modified cells and remove cells with undesirable insertions. The efficiency of CRISPR/Cas9-mediated HDR was increased up to 90% via G418 enrichment. We have successfully inserted eGFP under control of the chicken GAPDH promoter. Conclusions: The approach can be used further to insert genes of interest under control of tissue-specific promoters in primordial germ cells in order to produce genetically modified birds with useful for biotechnological purposes features.

Molecular modeling of immersion optical clearing of biological tissues.

The interaction of six low-molecular tissue-clearing agents (1,2 and 1,3-propanediol, ethylene glycol, glycerol, xylitol, sorbitol) with the collagen mimetic peptide (GPH)3 was studied by applying the methods of classical molecular dynamics (GROMACS), molecular docking (AutoDock Vina) and quantum chemistry (PM6 and B3LYP). The spatial configurations of intermolecular complexes were determined and interaction energies calculated. The dependence of the volume occupied by the collagen peptide on the clearing agent concentration in an aqueous solution was calculated. This dependence is not linear, and has a maximum for almost all the agents in the study. The correlations between the optical clearing potential and intermolecular interactions parameters, such as the time of an agent being in a hydrogen-bonded state, and the relative probability of formation of double hydrogen bonds and interaction energies, were determined. Using the correlations determined, we predicted the numeric value of the optical clearing potential of dextrose molecules in rat skin, which correlates with experimental data. A molecular mechanism of tissue optical clearing within the post-diffusion stage is suggested. Graphical abstract The molecular modeling of the interaction between clearing agents and collagen.

Biochemical adaptations to dive-derived hypoxia/reoxygenation in semiaquatic rodents.

To meet the challenges presented by dive-derived hypoxia/reoxygenation transition, the aquatic mammals possess multi-level adaptations. However, the adjustments of the semiaquatic animals as modern analogs of evolutionary intermediates between ancestral terrestrial mammals and their fully aquatic descendants are still not fully elucidated. The aim of this study was to analyze the total lactate dehydrogenase (LDH) activity (in the lactate to pyruvate direction), the LDH patterns and the antioxidant defense in the tissues (heart, kidney, liver, lung, muscle, spleen) of semiaquatic rodents such as Eurasian beaver (Castor fiber), muskrat (Ondatra zibethicus) and nutria (Myocastor coypus). Samples from Wistar rat were used for comparison. Semiaquatic rodents had higher catalase activity compared to rats. The superoxide dismutase activity was higher and the catalase activity was lower in almost all tissues of muskrat than of both beaver and nutria. Comparing beaver and nutria, no significant differences in the antioxidant enzyme activities were found for the heart, kidney and liver. In beaver, most of the examined tissues (heart, kidney, lung and spleen) use lactate as preference to glucose as a substrate but in muskrat the heart, liver and skeletal muscle showed the increased LDH activity. Nutria had the unusual LDH properties that are needed to be further investigated. Our results suggest that beaver, nutria and muskrat have distinct mechanisms of adaptation to diving hypoxia/reoxygenation and support the hypothesis that semiaquatic mammals are the intermediate animals that help to define which potential selection factors and mechanical constraints may have directed the evolution of the aquatic forms.