H2S-Mediated Changes in Erythrocyte Volume: Role of Gardos Channels, Na+,K+,2Cl- Cotransport and Anion Exchanger.

The effect of H2S on changes in erythrocyte volume was studied by spectrophotometrical and potentiometric methods. It was found that H2S donor NaHS (2.5, 10, and 100 µM) induced an increase in erythrocyte volume in heterosmotic media. Activation of Gardos channels with A23187 or ascorbate-phenazine methosulfate system causes erythrocyte shrinkage and hyperpolarization of their membrane, while addition of NaHS restored erythrocyte volume. The decrease in erythrocyte volume upon blockade of Na+,K+,2Cl- cotransporter (bumetanide) or anion exchanger (SITS) was abolished by H2S donor NaHS, which attested to an important role of these transporters and chlorine conductivity of the membrane in the maintenance of the homeostasis of blood cells.

Role of Carbon Monoxide in the Mechanisms of Action of Extracellular ATP on Contractile Activity of Vascular Smooth Muscle Cells.

We studied the role of carbon monoxide (CO) in the effect of P2X and P2Y receptor agonist ATP on the tone of rat aorta segments with intact endothelium. ATP (1-1000 µM) and P2X receptor agonist α,ß-MeATP (100 µM) relaxed segments precontracted with phenylephrine (10 µM), while UTP (100-1000 µM) increased the amplitude of phenylephrine-induced contraction. The relaxing effect of ATP was enhanced by CORM II (100 µM), NO synthase inhibitor L-NAME, and guanylate cyclase inhibitor ODQ and attenuated by ZnPP IX (100 µM). The constrictive effect of UTP was weakened by CORM II (100 µM), but was not changed by ZnPP IX (100 µM). ZnPP IX (100 µM) weakened the relaxation response to α,ß-MeATP. Thus, ATP involves the CO-dependent signaling cascade through P2X receptors.

Cytokine Profiling of Subclinical Tick-Borne Infections in Humans.

Over many years, tick-borne infections remain one of the most serious threats to human health worldwide. The immune response to these infections in a human after confirmed bite by an infected carrier at the early stages of infection in the absence of clinical symptoms can be the first indicator of the presence of the infectious agent in the body. During viral infection, the concentration of IL-1α, IL-8, IL-10, IL-17A, and IFNγ increases; superoxide dismutase also increases, in contrast to bacterial infections. A slight decrease in the concentration is observed only for receptor antagonist IL-1Ra. During the infection caused by bacterial pathogens, very similar profiles of the innate human immune response are observed: activation of IL-1α, IL-8, and IFNα and suppression of superoxide dismutase, IL-1Ra, and IL-17A production. It has been demonstrated, that the immune response is triggered immediately after infection, and changes in the concentration of the main cytokines in the blood plasma can be detected as early as on days 2-5 after tick bite. These results can be useful in developing new methods of emergency diagnosis and prevention of tick-borne infections.

[Markers of gluten intolerance in children with autism spectrum disorders and Down'syndrome]. / Markery neperenosimosti gliutena u detei s rasstroistvami autisticheskogo spektra i sindromom Dauna.

AIM: To study serological and genetic markers of gluten intolerance in children and teenagers with autism spectrum disorders (ASD) and Down's syndrome (DS). MATERIAL AND METHODS: Thirty-three children with ASD (group 1) and 8 with DS (group 2), aged from 2.5 to 15 years, were examined. There were 27 boys and 6 girls in group1, 5 boys and 3 girls in group 2. Most of the children were on a regular diet and only 4 children with ASD kept gluten-free diet (GFD). Using ELI method antibodies to gliadin IgG (AntiGliadin IgG), antibodies to deamidated peptides of gliadin IgA (AntiDGP IgA), immunoglobulin A (IgA) were identified. Haplotypes HLA-DQ2 and DQ8 were determined using PCR. RESULTS: AntiGliadin IgG were identified in 12.1% (4) patients of group 1, with the exception of patients on GFD in 13.8%, and in 50% patients of group 2. One child with ASD had selective IgA deficiency. Haplotypes predisposing to celiac disease had 41.9% of patients of group 1 and 37.5% of patients of group 2. In ASD, the distribution of genotypes was as follows: DQ2 (64.3%), DQ8 (28.6%), DQ2/DQ8 (7.1%,). In DS, all patients had haplotype DQ2. AntiDGP IgA were not identified in both groups. CONCLUSION: The predominant form of gluten intolerance in children with ASD and DS is sensitivity to gluten, which can be identified in 40-50% of patients. Celiac disease, an autoimmune form of gluten intolerance, can be diagnosed in single cases, although predisposition to it is identified in 41.9% - 37.5% patients with ASD and DS, respectively. Before the start of GFD, laboratory tests should be made to identify forms of gluten intolerance and the use of GFD.

Molecular Mechanisms of Action of Gas Transmitters NO, CO and H2S in Smooth Muscle Cells and Effect of NO-generating Compounds (Nitrates and Nitrites) on Average Life Expectancy.

Gaseous signaling molecules (gas transmitters) take an especial position among the numerous signaling molecules involved in the regulation of both intracellular processes that occur in different types of cells and cell-cell interactions. At present time, gas transmitters include three molecules whose enzymatic systems of synthesis and degradation, physiological action and intracellular effectors, the change of which under the action of gas transmitters may result in physiological and/or pathophysiological effects are well- determined. These molecules include nitrogen oxide (NO), carbon monoxide (CO) and hydrogen sulfide (H2S). They are involved in the regulation of functions of various organs and systems of the human body, including the circulatory system. Interaction of NO, CO and H2S with various enzymatic and structural components of endothelial and, especially, smooth muscle cells has a significant impact on vascular tone and blood pressure. Furthermore, the crossing of NO-, CO- and H2S-mediated signaling pathways at common effectors and interaction with each other can determine the end, resulting functional response of the cell. The knowledge of the molecular targets of gas transmitters' action, the structure of the binding centers for gas transmitters and their interaction with each other may be essential in the development of methods of regulation of these signaling systems by targeted, directed action. This review summarizes the molecular mechanisms of the NO, CO and H2S interaction with the main targets, which carry out their regulatory effect on vascular smooth muscle cells. Also we describe here different ways of cross-regulation of NO-, CO- and H2S-dependent signaling pathways. We analyzed NO-synthase and nitrite reductase systems of nitric oxide cycle and discuss the nitrate-nitrite background of the existence of modern man, which can substantially modify the signaling system, the metabolism of virtually all cell ultrastructure of neurons, neuron-neuron and neuron-glial interactions and exerts its influence on socially significant diseases that can affect the quality and the average life expectancy.

Regulation of Contractile Responses of Vascular Smooth Muscle Cells under Conditions of Hypoxia-Reoxygenation.

We analyzed the effects of hypoxia and reoxygenation on changes in contractile activity in rat aortic smooth muscles. Both hypoxia and reoxygenation induced relaxation of smooth muscle cells precontracted with high-potassium Krebs solution (30 mM KCl) or α1-adrenoceptor agonist phenylephrine. Vasodilation resulted from enhancement of potassium permeability of smooth muscle cell membranes caused by activation of voltage-gated potassium channels (triggered by both precontracting agents) or by opening of ATP-sensitive potassium channels (phenylephrine). In isolated smooth muscle cells, both hypoxia and inhibition of Na+,K+-ATPase with ouabain led to depletion of intracellular store of macroergic substances, reduced potassium concentration, and elevated the content of sodium ions.

[Child and Adolescent Interepydemic Immunity to Influenza Viruses].

OBJECTIVE: Our aim was to evaluate the child and adolescent population immunity to influenza A viruses (IAV) and influenza B virus (IBV). METHODS: The concentration and specificity of antiviral antibodies was evaluated by hemagglutination inhibition assay (HAI) that was performed using commercial HAI diagnostic kits. RESULTS: The serum samples of 254 clinically healthy children and adolescents were examined in this study. 245 participants had the antibodies to IAV, 199--to IAV and IBV and only 4 children aged between 1 and 4 years and a 12-year-old boy had no immunity to IAVor IBV. The number of children with specific immunity increased in elder groups by 43% (from 81 to 116) for N0N1, and H3N2 subtypes and by 110% (from 38 to 80) for H1N1 subtype of IAV. In children younger than 4 years the titer of specific antibodies against the H1N1pdm09 was 1:210 and against the H3N2--1:270, whereas in adolescents of 10-14 years these figures were by 1.6 and 2.4 times lower (1:130 and 1:120) respectively. Antibodies to the subtypes H2N2 and H5N1 were not detected. CONCLUSION: The results indicate that 98.4% of child and adolescent population in interepidemic influenza season are immune to the various IAV (H3N2, H1N1, H0N1) as well as to the IBV. More than half of children and adolescents (57.4%) are immune to H1N1pdm09 subtype of IAV. The strength of immune response to the recent pathogens (H3N2 u H1N1pdm09) is higher in infants than in teenagers.

[Gas Signalling in Mammalian Cells].

At the end of the last century after the discovery of signaling functions of nitric oxide (NO, II), a new class of biologically active substances was admitted. It includes so-called gas transmitters acting as intercellular and intracellular regulators of different physiological functions. Currently, this class includes such gases as NO, carbon monoxide (CO) and hydrogen sulfide (H2S). It was found that these gases regulate not only functions of the. gastrointestinal tract and the cardiovascular system, where it has been determined initially, but also affect the function of the central and peripheral nervous.systems. Apparently, they constitute a single complex of gas transmitters, which easily penetrates through the membrane and regulates numerous enzymatic and non enzymatic cells reactions. This review presents the mechanisms of gas transmitters' influence on the electrical and contractile properties of smooth muscle cells (SMC) as a possible new ways to interact with the "classical" intracellular signaling cascades (Ca2+, cyclic nucleotides) and effectors systems. On account of their interactions the role of cyclic nucleotides and calcium ions in the implementation of the signal gas molecules functions is analyzed. We summarize the literature data and the results of our own research on the role of SMC membrane ion-transporting systems in myogenic effects of NO, CO and H2S and describe possible reasons of gas transmitters multidirectional influence on the excitation-contraction coupling in SMC.

[THE INFLUENCE OF HYDROGEN SULFIDE ON COLLAGEN-INDUCED AGGREGATION OF HUMAN PLATELETS].

Study the impact of hydrogen sulfide on collagen-induced platelet aggregation from healthy donors and patients with type 2 diabetes. In healthy individuals, in contrast to patients with type 2 diabetes, NaHS significantly inhibited platelet aggregation. Activators of cAMP signaling (forskolin and phosphodiesterase inhibitor) significantly reduced platelet aggregation in both groups of examinees. NO-synthase inhibitors increased platelet aggregation in healthy volunteers, but not in patients with type 2 diabetes. The presence of H2S donor did not alter the extent of platelet aggregation at high concentrations of cAMP or decreased production of nitric oxide. It is assumed that the antiplatelet effect of H2S is not associated with the effect on the signal system, mediated cAMP or nitric oxide. Change H2S-dependent regulation of platelet aggregation in patients with type 2 diabetes is caused by disorders have been reported with this disease: the increase of intracellular calcium ion concentration, oxidative damage to proteins, hyperhomocysteinemia, glycosylation of key proteins involved in this process.

[Postglacial migration and phenogeography of populations of the Scots pine (Pinus sylvestris L.) in the northeast of the Russian Plain].

The history, distribution routes, and phenogeographic structure of the Scots pine (Pinus sylvestris L.) in the northeast of the Russian Plain were studied on the basis of paleogeographic data and results of our own phenotypic and allozyme-genetic studies. It is assumed that, after the maximum Dnieper glaciation, P. sylvestris populations could successfully distribute to the northwest and north from the refugia of the South and Middle Urals as a result of seed dispersal by Belaya, Ufa, Chusovaya rivers (in Holocene, by Severnaya Dvina, Mezen', and Pechora rivers). On the basis of the hypothesis of "migration complexes" and the theory of hydrochory for coniferous species, a scheme of formation of a population structure of the Scots pine in the northeast of the Russian Plain is proposed.

[Genetic and phenotypic differentiation of Calluna vulgaris (L.) Hull in Pritobolie and Europe].

Geographic variation and differentiation of the chloroplast DNA haplotypes and morpho-anatomical leaf parameters were assessed in a number of eastern European groups of Calluna vulgaris (L.)Hull populations and in the Pritobolien group of populations of this species in Western Siberia, which have long been isolated from the European populations. Sharply pronounced genetic and phenotypic distances and their gradients between the Pritobolien and European population groups were revealed. These distances were many times higher than those between the relatively homogeneous eastern European groups. The data obtained generally supported the hypothesis on the phenogenetic divergence of the Pritobolien marginal populations of C. vulgaris from the European, probably at the subspecies level.

Adlayers of palladium particles and their aggregates on porous polypropylene hollow fiber membranes as hydrogenization contractors/reactors.

Principal approaches for the preparation of catalytic membrane reactors based on polymer membranes containing palladium nanoparticles and for the description of their characteristics are presented. The method for the development of adlayers composed of palladium nanoparticles and their aggregates on the surface of hydrophobic polypropylene porous hollow fiber membranes is proposed, and their comprehensive study is performed. Various regimes of the deposition of palladium on individual fibers and on membrane surface inside membrane modulus are considered. The sizes of primary Pd particles range from 10 to 500 nm, and dimensions of their aggregates vary from 200 nm to tens of microns. The sizes of primary particles in a free state and in their aggregates are estimated by the methods of X-ray analysis and scanning electron microscopy. The proposed approach is used for the preparation of catalytic membrane contactors/reactors for the removal of dissolved oxygen from water. In the systems under study, the limiting stage of dissolved oxygen removal is concerned with diffusion-controlled delivery of oxygen to the surface of catalytic particles.

[Genetic diagnostic technique for opisthorchiasis].

OBJECTIVE: To develop a test system for polymerase chain reaction (PCR) diagnosis of Opisthorchis felineus in human feces. Based on the Internet databases, the authors have developed oligonucleotide primers, as well as TaqMan tests (fluorescence label probes) specific to O. felineus genome to perform PCR with the real-time detection of a result. Conditions for PCR, including temperatures, the number of components of amplification, etc., were selected. Test systems on fecal samples from 254 subjects randomly selected and on those from 100 patients with verified opisthorchiasis, and on those from 100 healthy individuals were studies. The significance of the test systems was ascertained to be about 98%. Their sensitivity in determining O. felineus is about 10(3) copies of target DNA per ml. The efficiency of various techniques (coproovoscopy, enzyme immunoassay, PCR) was compared to detect Opisthorchis invasion.

[Nitric oxide metabolites and their significance in pathogenesis of bronchial asthma].

To study the possible potential of nitric oxide (NO) in the processes of atopic inflammation, the authors evaluated the intensity of nitrosyling and oxidative stresses in the bronchoalvelar lavage fluid and expired air condensate of patients with bronchial asthma (BA). Chronic inflammation was shown to result in an increase in the processes of nitration and oxidation in worsening BA and to reliably correlate with airway NO-producing functions, by explaining the pathological effects of NO due to the formation of 3-nitrotyrosine and malonic dialdehyde with the existing imbalance in NO metabolism, by intensifying nitrosylating stress. In the authors' opinion, nitrosothiols that are required as a NO donor may be rapidly destroyed or virtually do not form so the peroxynitrite-nitrosothiol ratio may predetermine the final effects of NO.

[Phenazine methosulfate system-induced membrane hyperpolarization in the human erythrocytes].

Erythrocyte membrane potential was recorded via measurement of pH of the incubation medium in presence ofprothonophore. The increase of intracellular calcium concentration in presence of calcium ionophore A23187 and addition of the artificial redox-system ascorbate-phenazine methosulfate led to membrane hyperpolarization due to opening of Ca(2+)-activated potassium channels that are regulated by multiple signaling pathways. The opening of the Ca(2+)-activated potassium channels in presence of artificial redox-system ascorbate-phenazine methosulfate is mediated at least by two mechanisms including an increase in affinity of channels to calcium ions and involvement of the protein SH-groups and the components of the respiratory circuit which have beer found in erythrocyte membrane.

Effect of insulin on Ca(2+)-dependent hyperpolarization in erythrocytes from healthy donors and patients with type 2 diabetes mellitus accompanied by arterial hypertension.

Insulin decreased A23187-induced hyperpolarization of the erythrocyte membrane in healthy donors. These data indicate that insulin plays a role in the regulation of Ca(2+)-activated potassium channels in human erythrocytes. However, insulin had little effect on hyperpolarization response of cells induced by artificial ascorbate--phenazine methosulfate donor-acceptor system. Addition of insulin to cell suspension from patients with type 2 diabetes mellitus did not modulate hyperpolarization of the erythrocyte membrane induced by A23187 or ascorbate-phenazine methosulfate, which reflects impairment of regulatory mechanisms for Ca(2+)-activated potassium channels in erythrocytes.

Volume-dependent regulation of Ca2+-activated potassium channels in erythrocytes from healthy donors and patients with type II diabetes mellitus aggravated by arterial hypertension.

Increase in intracellular Ca2+ concentration caused by calcium ionophore A23187 or ascorbate+phenazine methosulphate electron donor system added to erythrocyte suspension induced similar shifts in erythrocyte membrane potential. These processes are most likely mediated by Ca2+-activated potassium channels. Changes in the osmolarity of the incubation medium produced opposite effects on membrane hyperpolarization induced by A23187 or ascorbate+phenazine methosulphate in erythrocyte isolated from healthy donors, which attests to the existence of different mechanisms of regulation of Ca2+-activated potassium channels. There was no difference in the volume-dependent changes of potassium permeability in cells from patients with type II diabetes mellitus combined with arterial hypertension induced by application A23187 or electron-donor system.