A sensitive and less cytotoxic assay for identification of proliferating T cells based on bioorthogonally-functionalized uridine analogue.

Quantitative detection of T cell proliferation is an important readout in immunology research, as it is one of the hallmarks of T cell activation. Fluorescence-based methods for T cell proliferation mostly rely on the usage of probes that non-specifically conjugate to free primary amine groups in cells. Each cell division then results in a two-fold dilution of the probes which is detectable with flow cytometry. However, questions have been raised about cytotoxicity of these dilution-based T cell proliferation probes and they potentially affect T cell activation. An alternative assay relies on the incorporation of the uridine analog BrdU in the DNA of dividing T cells that can be detected with an antibody, but this requires harsh fixation and denaturation conditions. Recently, a new assay for detection of cell proliferation has been developed, based on the incorporation of the bioorthogonally-functionalized uridine analog 5-ethynyl-2'-deoxyuridine (EdU). Goal of this study was to compare the sensitivity and cytotoxicity of the EdU assay with a widely-used dilution-based T cell proliferation probe, CellTrace Far Red. We found that, compared to the dilution-based probe, the EdU-based assay better preserves T cell viability, is more sensitive for detecting T cell proliferation, and allows for better discernable interferon gamma responses.

A Method of Ground Simulation of Physiological Effects of Hypogravity on Humans.

A novel method of ground simulation in humans of physiological effects induced by the stay on the surface of celestial bodies with hypogravity was developed and successfully tested. This method is based on the change of gravity force angle, which decreases the gravitational component of the blood hydrostatic pressure characteristic of human vertical posture on the Earth and the load-weight onto the locomotor apparatus to the lower values expected at celestial bodies with hypogravity. The methodological requirements for ground simulation of the physiological effects of lunar gravity on human body are specified and substantiated by theoretical calculations. The experimental study revealed redistribution of liquid media in the human organism, functional changes in the cardiorespiratory system, and a decrease in the load-weight applied to the locomotor apparatus.

[Influence of simulated microgravity on the threshold of pain sensitivity in humans with single dose of ketorolac].

The data supporting the influence of simulated microgravity effects on pain sensitivity were obtained in the series of experiments involving human. In conditions of antiorthostatic hypokinesia (ANOH) and <> immersion revealed no reduction in pain sensitivity in the morning, which is typical for normal conditions. Ketorolac has no effect on pain sensitivity, when determining the pain threshold (PT) by method of thermoalgometry. However, the conditions of simulated microgravity substantially alter the pharmacokinetics of ketorolac, increasing the rate of absorption of the drug and reduce its relative bioavailability and retention time in the blood plasma. This may require changes in pain therapy schemes in space flight.

[Features of the pharmacokinetics of ceftriaxone during the antiorthostatic hypokinesia in rats].

This study was designed to investigate the effects of simulated microgravity on the plasma kinetics of ceftriaxone in rats. Comparative analysis of pharmacokinetic parameters in control and experimental groups has revealed the influence of antiorthostatic hypokinesia (ANOH) on the pharmacokinetics of ceftriaxone. Decreased level of maximum plasma concentration (C(max)) and bioavailability of ceftriaxone after single intramuscular (IM) injection were revealed in rats with antiorthostatic hypokinesia. The rate of absorption of the drug was higher respectively.

Polyphosphates and exopolyphosphatase activities in the yeast Saccharomyces cerevisiae under overexpression of homologous and heterologous PPN1 genes.

The role of exopolyphosphatase PPN1 in polyphosphate metabolism in fungi has been studied in strains of Saccharomyces cerevisiae transformed by the yeast PPN1 gene and its ortholog of the fungus Acremonium chrysogenum producing cephalosporin C. The PPN1 genes were expressed under a strong constitutive promoter of the gene of glycerol aldehyde-triphosphate dehydrogenase of S. cerevisiae in the vector pMB1. The yeast strain with inactivated PPN1 gene was transformed by the above vectors containing the PPN1 genes of S. cerevisiae and A. chrysogenum. Exopolyphosphatase activity in the transformant with the yeast PPN1 increased 28- and 11-fold compared to the mutant and parent PPN1 strains. The amount of polyphosphate in this transformant decreased threefold. Neither the increase in exopolyphosphatase activity nor the decrease in polyphosphate content was observed in the transformant with the orthologous PPN1 gene of A. chrysogenum, suggesting the absence of the active form of PPN1 in this transformant.

[Biochemical aspects of the influence of antiorthostatic hypokinesia on the experimental peritonitis].

Eighty white rats were divided into 4 groups: (1) vivarium control, (2) antiorthostatic hypokinesia (AH), (3) peritonitis alone, and (4) AH with peritonitis. Effects of AH were achieved by putting rats on a special stand for a period of 14 days, followed by the formation of peritonitis. After that biochemical parameters of blood samples have been investigated. Combination effects of microgravity and peritonitis is unidirectional and have mutual weights. As illustrated in some cases two-fold increase in the level of the studied parameters in comparison with the control and vivarium peritonitis. Comparative assessment of peritonitis severity revealed that simulated effects of microgravity turn the peritonitis into more complicated forms.

[Physiological effects of microgravity as risk factors of diseases during space flight].

This paper considers to physiological effects of microgravity affecting different organs systems such as cardiovascular, neurohumoral, immune, and musculoskeletal systems. These physiologic changes suggest adverse influence with pathological processes during space flight and associated with high risk of morbidity of cosmonauts.

[Formation of spatial structure of RNA molecules].

In this review we consider several experimental and theoretical approaches for investigation of RNA folding and determination of nucleotides that play an important role upon folding of such molecules as tRNA and several classes of ribozymes. It has been shown that nucleotides in the D- and T-loop regions are the last to be involved in tRNA structure or they are not included in the folding nucleus of tRNA. Using the specially elaborated method SHAPE it has been demonstrated that the model of hierarchical folding which was recognized for a long time is not correct for tRNA folding. In the second part of the given review the algorithms and programs used for the prediction of secondary structures of RNA as well as for modeling of RNA folding are considered.

[Change in sensitivity of the central respiration mechanism in the 21-hour bedrest conditions].

The investigation was aimed at studying the mechanisms for change in the respiration center sensitivity in consequence of 21-hr bed rest with head-end tilted at -15 degrees combined with liquid loss (lasix, 20 ml) and recovery (IV infucol and glucose). Time of maximal breath-holding, capillary and venous O2 and CO2 pressure values were measured in the baseline data collection period, during and shortly after BR. Data analysis showed that extension of the maximal breath-holding time both during inspiration and expiration was statistically significant in the initial 10 minutes of tilting. Comparison of the breath-holding test data between the experimental series demonstrated that infusion of equally glucose and infucol did not affect voluntary apnea during inspiration or expiration. From BR hour 17, partial pressure of venous O2 showed a significant rise, while venous CO2 pressure decreased, also significantly. It is hypothesized that degradation of the respiration center sensitivity was connected most likely with blood pooling in the upper body and altered pressure on the baroreceptors.

[Structural and functional features of the rats' prefrontal cortex following 14-day suspension].

Effects of 14-d suspension on the processes of rats' nervous cells conjugation in the brain cognitive cortex were studied. Signs of cognitive functions degradation and decrease of the number of conjugated nervous cells were found in the experimental group after suspension. This evidence are qualified as a negative effect of suspension on memory and physiological regeneration of neurons.

Prediction of folding nuclei in tRNA molecules.

Prediction of folding nuclei in RNA molecules allows one to look in a new way at the problem of possible RNA base sequence folding and at problems associated with incorrect RNA folding, as well as at RNA structure stability. We have chosen a model and energy parameters for description of RNA structure. The algorithm for studying processes including protein folding/unfolding was successfully applied to calculations on tRNA. Four tRNA molecules were considered whose structures were obtained in the free state (tRNA(Phe), tRNA(Asp), tRNA(fMet), and tRNA(Lys)). The calculated Φ-values for tRNA molecules correlate with experimental data showing that nucleotide residues in the D and T hairpin regions are involved in tRNA structure last, or more exactly, they are not included in the tRNA folding nucleus. High Φ-values in the anticodon hairpin region show that the nucleus of tRNA folding is localized just in that place.

[Effects of dehydration and infusion of blood substitutes on the hemostasis system in an experiment with 21-hour bed rest].

Eight volunteered male test-subjects (20-40 y.o.) took part in an experiment with 21-hr bed rest aimed to investigate trends in the hemostasis parameters (activated partial thromboplastin time, prothrombin time, thrombin time, fibrinogen, plasminogen, D-dimers, soluble fibrin-monomer complexes (SFMC), as well as antithrombin III (ATIII) and protein C (antiplasmin) activities) after diuretics-induced dehydration followed by intravenous infusion of colloid and crystalloid solutions. It was shown that noncompensible dehydration during bed rest does not lead to noteworthy alterations in the procoagulating component of hemostasis. Intravenous infusion of blood substituting sterofundin (crystalloid) and venofundin (colloid) to the bedrested human subjects accelerated coagulation within the physiological norm. In all series of the experiment ATIII was suppressed and plasminogen content went down causing growth in SFMC. Fibrinogen concentration remained essentially unchanged pointing to the absence of acute reaction of organism to the experimental conditions.

[Methods of pain sensitivity evaluation in the experiment and in clinical practice].

In the article different methods of pain sensitivity evaluation are covered both in the experiment and in clinical practice, positive and negative sides are determined of different methods of pain sensitivity threshold determination.

[Testing and selection of equipment to simulate in-flight telemedicine-assisted medical examinations and researches on the ISS Russian segment].

Analysis of equipment and systems used in telemedicine consultation was fulfilled from the perspective of integration into the ISS RS.

[Nociceptive sensitivity and environmental factors]. / Notsitseptivnaia chuvstvitel'nost' i faktory okruzhaiushchei sredy.

The authors dwell upon the structure and functioning of the nociceptive system and modification of pain sensitivity by ambient factors like air temperature and atmospheric pressure.

[Peculiarities of anesthesia and systemic hemodynamics in experiment with 30 days head-down immobilization of monkeys]. / Osobennosti techeniia anestezii i sostoianiia sistemnoi gemodinamiki v éksperimente s 30-sutochnoi antiortostaticheskoi gipokineziei u obezi'ian.

Two series of surgery (biopsy) were performed before and after 30-d head-down immobilization (HDI) of monkeys. The animals were anaesthetized by inhaled isoflurane. Effects of immobilization on systemic hemodynamics and the progress of anesthesia were evaluated. The experiment showed that HDI reduces systemic arterial pressure and slightly increases heart rate (HR) irrespective of the use of anesthesia. However, comparison of systemic hemodynamics parameters before and after the 30-d immobilization also revealed a drop in systemic arterial pressure, both systolic and, to a less degree, diastolic but not a significant rise in HR during the second surgical operation.

[Waves on the cell surface]. / Volny na kletochnoi poverkhnosti.

A new theoretical model is presented which describes dynamics of interaction between the plasmic membrane, cell cortex and cytoplasmic liquid. Analysis of the equation system describing this interaction has shown that besides the uniform solution (corresponding to a flat cell surface) there exists the stable autowave solution (corresponding to standing or running harmonic waves along the cell surface). The latter solution can be related to the experimentally observed waves moving along the cell surface.

[The computer at the pathologist's laboratory]. / Komp'iuter v patologoanatomicheskoi laboratorii.

Computers are coming into practical use by pathologists. The computer hardware and software are briefly outlined. The software is discussed while drawing up protocols on the examination of biopsies, operative materials and autopsies. A special attention is given to the SNOP and SNOMED coding of diagnoses. How to operate on a computer is described in the paper. The latter provides evidence for applying the computer to the formulation of laboratory documents.