Spectroscopic analysis of metabolic profile in patients with relapsed multiple sclerosis

Introduction. Managing patients with relapsing-remitting multiple sclerosis (RMS) remains a pressing issue. Objective. To detect the reversible metabolic changes of the brain matter in patients with clinically exacerbated RMS and to follow them up after intravenous glucocorticoid (IVGC) treatment. Materials and methods. Neurological examination and neuroimaging in the RMS patients included expanded disability status scale (EDSS) scoring, conventional brain magnetic resonance imaging (MRI), and proton nuclear magnetic resonance spectroscopy (1H-NMR spectroscopy) before and after IVGC treatment. Multivoxel 1H-NMR spectroscopy was used to assess metabolism in the centra semiovale and cingulate gyri. Results. Based on the multivoxel 1H-NMR spectroscopy, relative metabolite concentrations in the grey and white matter statistically differed within the study cohort before and after the IVGC treatment. The N-acetylaspartate/choline ratio significantly recovered and the choline/creatine ratio decreased in the anterior cingulate gyri in 27% of patients. The brainstem function score significantly improved in the metabolic response group as compared to the non-metabolic response group. Conclusion. We should study the potential predictors of RMS activity and the IVGC response to select the RMS relapses when pulse-therapy with IVGCs is definitely indicated. Spectroscopy may reveal RMS pathogenesis variability earlier than conventional MRI

Mitochondria Express α7 Nicotinic Acetylcholine Receptors to Regulate Ca2+ Accumulation and Cytochrome c Release: Study on Isolated Mitochondria

Nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that mediate synaptic transmission in the muscle and autonomic ganglia and regulate transmitter release in the brain. The nAChRs composed of α7 subunits are also expressed in non-excitable cells to regulate cell survival and proliferation. Up to now, functional α7 nAChRs were found exclusively on the cell plasma membrane. Here we show that they are expressed in mitochondria and regulate early pro-apoptotic events like cytochrome c release. The binding of α7-specific antibody with mouse liver mitochondria was revealed by electron microscopy. Outer membranes of mitochondria from the wild-type and β2−/− but not α7−/− mice bound α7 nAChR-specific antibody and toxins: FITC-labeled α-cobratoxin or Alexa 555-labeled α-bungarotoxin. α7 nAChR agonists (1 µM acetylcholine, 10 µM choline or 30 nM PNU-282987) impaired intramitochondrial Ca2+ accumulation and significantly decreased cytochrome c release stimulated with either 90 µM CaCl2 or 0.5 mM H2O2. α7-specific antagonist methyllicaconitine (50 nM) did not affect Ca2+ accumulation in mitochondria but attenuated the effects of agonists on cytochrome c release. Inhibitor of voltage-dependent anion channel (VDAC) 4,4′-diisothio-cyano-2,2′-stilbene disulfonic acid (0.5 µM) decreased cytochrome c release stimulated with apoptogens similarly to α7 nAChR agonists, and VDAC was co-captured with the α7 nAChR from mitochondria outer membrane preparation in both direct and reverse sandwich ELISA. It is concluded that α7 nAChRs are expressed in mitochondria outer membrane to regulate the VDAC-mediated Ca2+ transport and mitochondrial permeability transition

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Energy security is an important component of national security

The article deals with the issues of energy security as an important component of the national security of the country. There is growing uncertainty about safe access to reliable energy supplies at reasonable prices

Changing national health values of Russians: Perception of physical culture and sports

The publication is aimed at studying the attitude of middle-aged and older Russians to physical culture and sports. The scope of the results embraces studying the dynamics of perception of physical culture and sports in the minds of middle-aged and older Russians. The research subject is studying the dynamic changes in the system of value consciousness of Russians. The method of obtaining empirical data includes focused online interviews, traditional analysis of documents, including legislative acts of the Russian Federation. Conducting focused online interviews with respondents from Nizhny Novgorod and Volgograd is due to the conditions of self-isolation of Russians during the study period. The result of the study is presented as a description of the specifics of the perception of physical culture and sports by middle-aged and older Russians. The scientific novelty of the publication lies in the author's understanding of the influence of self-isolation on changes in the values of Russians in the perception of physical culture and sports worthiness by middle-aged and older people in Russian regions. During the interviewing online focus groups, respondents' suggestions on how to encourage middle-aged and older Russians to engage in physical culture and sports were revealed

The historical consciousness of student youth and evaluation of the events of the Great Patriotic War

It is not the first year that the information space has been marked by trends in the revision of evaluations of the events of the Great Patriotic War. Student youth are actively and rapidly developing, adapting to the new social realities. That is why the issues of preserving historical memory and developing the historical consciousness of contemporary young people have become so relevant. The present work is based on the analysis of the survey results of the 1st and 2nd-year students conducted at the State University of Management (SUM, Moscow, Russia), and is a continuation of the research devoted to the evaluation of the end results of the Great Patriotic War in the perception of University students. An attempt was made to determine the level of students’ knowledge about the main facts of the Great Patriotic War. Within the framework of this study, the results of similar works by domestic and foreign authors were considered. As a rule, in Russia, the problem of awareness of the importance of the Great Patriotic War is emphasized in terms of its results, including the perception of these issues by student youth. At the same time, several foreign authors put forward the issue of reevaluating the results of the Second World War and make it with clear politicization of the proposed conclusions, primarily, in determining the historical responsibility for war unleashing. The authors of the present study propose to strengthen the role of archival documents, memoirs, chronicles, and testimonies of direct participants and victims of the wartime events in military-historical education, which should not allow distorting the historical feat of the USSR and its decisive contribution to the defeat of Nazi Germany

Engineering of Thermal Stability in a Cold-Active Oligo-1,6-Glucosidase from Exiguobacterium sibiricum with Unusual Amino Acid Content

A gene coding for a novel putative amylase, oligo-1,6-glucosidase from a psychrotrophic bacterium Exiguobacterium sibiricum from Siberian permafrost soil was cloned and expressed in Escherichia coli. The amino acid sequence of the predicted protein EsOgl and its 3D model displayed several features characteristic for the cold-active enzymes while possessing an unusually high number of proline residues in the loops—a typical feature of thermophilic enzymes. The activity of the purified recombinant protein was tested with p-nitrophenyl α-D-glucopyranoside as a substrate. The enzyme displayed a plateau-shaped temperature-activity profile with the optimum at 25 °C and a pronounced activity at low temperatures (50% of maximum activity at 5 °C). To improve the thermal stability at temperatures above 40 °C, we have introduced proline residues into four positions of EsOgl by site-directed mutagenesis according to “the proline rule”. Two of the mutants, S130P and A109P demonstrated a three- and two-fold increased half-life at 45 °C. Moreover, S130P mutation led to a 60% increase in the catalytic rate constant. Combining the mutations resulted in a further increase in stability transforming the temperature-activity profile to a typical mesophilic pattern. In the most thermostable variant A109P/S130P/E176P, the half-life at 45 °C was increased from 11 min (wild-type) to 129 min

Combining mKate2-Kv1.3 Channel and Atto488-Hongotoxin for the Studies of Peptide Pore Blockers on Living Eukaryotic Cells

The voltage-gated potassium Kv1.3 channel is an essential component of vital cellular processes which is also involved in the pathogenesis of some autoimmune, neuroinflammatory and oncological diseases. Pore blockers of the Kv1.3 channel are considered as potential drugs and are used to study Kv1 channels’ structure and functions. Screening and study of the blockers require the assessment of their ability to bind the channel. Expanding the variety of methods used for this, we report on the development of the fluorescent competitive binding assay for measuring affinities of pore blockers to Kv1.3 at the membrane of mammalian cells. The assay constituents are hongotoxin 1 conjugated with Atto488, fluorescent mKate2-tagged Kv1.3 channel, which was designed to improve membrane expression of the channel in mammalian cells, confocal microscopy, and a special protocol of image processing. The assay is implemented in the “mix and measure”, format and allows the screening of Kv1.3 blockers, such as peptide toxins, that bind to the extracellular vestibule of the K+-conducting pore, and analyzing their affinity

GFP–Margatoxin, a Genetically Encoded Fluorescent Ligand to Probe Affinity of Kv1.3 Channel Blockers

Peptide pore blockers and their fluorescent derivatives are useful molecular probes to study the structure and functions of the voltage-gated potassium Kv1.3 channel, which is considered as a pharmacological target in the treatment of autoimmune and neurological disorders. We present Kv1.3 fluorescent ligand, GFP–MgTx, constructed on the basis of green fluorescent protein (GFP) and margatoxin (MgTx), the peptide, which is widely used in physiological studies of Kv1.3. Expression of the fluorescent ligand in E. coli cells resulted in correctly folded and functionally active GFP–MgTx with a yield of 30 mg per 1 L of culture. Complex of GFP–MgTx with the Kv1.3 binding site is reported to have the dissociation constant of 11 ± 2 nM. GFP–MgTx as a component of an analytical system based on the hybrid KcsA–Kv1.3 channel is shown to be applicable to recognize Kv1.3 pore blockers of peptide origin and to evaluate their affinities to Kv1.3. GFP–MgTx can be used in screening and pre-selection of Kv1.3 channel blockers as potential drug candidates