Vol 163, No 1 (2017)

The necessity of studying the Bernstein effect “repetition without repetition” is dictated by the absence of quantitative description of this effect and models that could describe such unique phenomena as repeated limb movements in a person in various mental states. In 30 nominally healthy volunteers (15 men, 15 women aged 24-25 years), tappingrams and tremorograms were recorded using an eddy current sensor with an oscillatory circuit (1 MHz) and an amplifier with recording frequency 0-1000 Hz and minimum amplitude of 0.01 mm. The results were recorded as files, processed as matrices of paired comparisons of samples, the number of matching sample pairs was determined (significance level p>0.05), and phase trajectories of finger movement were plotted. The effect was observed for both tapping and tremor and it is advisable to calculate the parameters of quasi-attractors that changed upon shifts in homeostasis.

Neuronal NO synthase blocker 7-nitroindazole suppressed bicarbonate secretion in rat gastric mucosa induced by mild local irritation with 1 M NaCl (pH 2.0). Non-selective blocker of neuronal and endothelial synthases, Nω-nitro-L-arginine (L-NNA), did not affect HCO₃^– production, but inhibited secretion after pretreatment with omeprazole. Non-selective cyclooxygenase blocker indomethacin inhibited HCO₃^– production under conditions of normal synthase activity and in the presence of L-NNA, but was ineffective when co-administered with 7-nitroindazole. It was concluded that neuronal and endothelial synthases are involved in different mechanisms of regulation of HCO₃^– secretion in the gastric mucosa induced by mild irritation. Activation of neuronal synthase stimulated HCO₃^– production, which is mediated mainly through activation of cyclooxygenase. Theoretically, activation of endothelial synthase should suppress HCO₃^– production. The effect of endothelial synthase depends on acid secretion in the stomach and bicarbonate concentration in the submucosa, as it was demonstrated in experiments with intravenous NaHCO₃ infusion.

We studied the role of cyclic nucleotides in the influence of hydrogen sulfide (H₂S) donor, sodium hydrosulfide (NaHS, 200 μM), on motor activity of rat jejunum. NaHS reduced spontaneous and carbachol-induced contractions of rat jejunum segment, which suggests that H₂S can act through mechanisms involving muscarinic receptor activation. Against the background of a membrane-penetrating non-hydrolyzable cAMP analogue or under conditions of adenylate cyclase blockade, the inhibitory effect of NaHS on the carbachol-induced contractions was maintained. Against the background of elevated cGMP concentration or guanylate cyclase inhibition, the reduction of carbachol-induced contractions upon exposure to NaHS was less pronounced than in control. It was hypothesized that H₂S induces relaxation of carbachol-induced jejunum contractions, affecting protein kinase G targets or activating cGMP synthesis.

In Wistar rats, stress was modeled by 24-h immobilization in a supine posture and stress-induced damage to the heart was assessed by accumulation of ^99mTc-pyrophosphate in the myocardium. The intensity of stress reaction was measured by serum levels of cortisol and insulin. Both stressinduced damage to the heart and intensity of stress reaction were examined under control conditions and in rats treated with opioid receptor antagonists naltrexone, methylnaltrexone bromide, MR2266, and ICI174.864. Activation of central μ-opioid receptors with endogenous opioids aggravated stress-induced cardiomyopathy, while stimulation of peripheral μ-opioid receptors produced a cardioprotective effect. The stress-induced damage to the heart was not directly related to up-regulation of cortisol secretion in response to 24-h immobilization. Blockade of the central opioid receptors promoted a decrease in cortisol level in stressed rats.

We studied heart rate variability and β-adrenergic responsiveness of erythrocytes and changes in these parameters in response to single administration of β-adrenoblocker propranolol (2 mg/kg) in outbred male rats against the background of activation of the noradrenergic, serotonergic, and dopaminergic neurotransmitter systems achieved by 4-fold injections maprotiline (10 mg/kg), 5-hydroxytryptophan (50 mg/kg) combined with fluoxetine (3 mg/kg), and L-DOPA (20 mg/kg) with amantadine (20 mg/kg), respectively. Stimulation of the noradrenergic system moderately enhanced the heart rhythm rigidity and β-adrenergic responsiveness of erythrocytes. In addition, it markedly augmented the moderating effect of subsequently administered propranolol on LF and VLF components in the heart rate variability and reversed the effect of propranolol on β-adrenergic responsiveness of erythrocytes. Stimulation of the serotonergic system dramatically decreased all components in the heart rate variability and pronouncedly enhanced β-adrenergic responsiveness of erythrocytes. Subsequent injection of propranolol slightly restored all components in the heart rate variability and decreased β-adrenergic responsiveness of erythrocytes to the control level. Stimulation of the dopaminergic system made the heart rate more rigid due to decrease of all components in the heart rate variability; in addition, it slightly but significantly enhanced β-adrenergic responsiveness of erythrocytes. Subsequent injection of propranolol produced no significant effects on all components in the heart rate variability and on β-adrenergic responsiveness of erythrocytes. Stimulation of noradrenergic, serotonergic, and dopaminergic neurotransmitter systems produced unidirectional and consorted effects on heart rate variability and β-adrenergic responsiveness of erythrocytes, although the magnitudes of these effects were different. Probably, the changes in the heart rate variability in rats with stimulated neurotransmitter systems results from modification of the cellular sensitivity in peripheral organs to adrenergic influences. However, the differences in the reactions to β-adrenoblocker attest to specificity of the mechanisms underlying the changes in membrane reception and adrenergic pathways in every experimental model employed in this study.

Experimental data on the involvement of the blood anticoagulant system components (heparin complex compounds, short regulatory peptides) in fibrin formation processes are presented. Inhibition of thrombin activity and interactions of short glyproline peptides and their heparin complexes with fibrin monomer molecules are demonstrated. Peptides and their heparin complexes prevent the formation of primary polymeric fibrin and exhibit fibrin-depolymerizing activity on the formed polymeric fibrin. The mechanisms underlying the effects of heparin complexes on degradation of fibrin not stabilized with factor XIIIa is discussed on the basis of the results of spectral analysis and coagulation tests.

The effects of dihydroquercetin (50 mg/kg intragastrically daily for 6 weeks) on the density of capillary network (mean number of capillaries per mm²), mean capillary diameter, structure of capillary network, capillary diameter distribution (<3, 3-5, 5-7, and 7-9 μ), and local cerebral blood flow (by laser Doppler) in the visual cortex were studied in SHR rats during the development of arterial hypertension (from the 6th to the 12th week of life). Normally, the systolic and diastolic BP progressively increased in SHR rats during this period. Dihydroquercetin did not affect the development of arterial hypertension. At the same time, the drug significantly increased the mean diameter of capillaries (by 11%), capillary network density (by 23%), and in the percentage of capillaries with a diameter of 3-9 μ (passable for erythrocytes; by 42%). Positive effects of dihydroquercetin on the structure of microcirculatory bed improved microcirculation: local cerebral blood flow in the visual cortex of SHR rats was significantly higher (by 36%) than in rats receiving no flavonoid and close to the value in Wistar-Kyoto rats. Dihydroquercetin improved microvascularization and microcirculation in the cerebral cortex of SHR rats during the formation of arterial hypertension.

Numerical density of vessels in axillary lymph node structures in breast cancer patients was studied by light microscopy with antibodies to CD34. The correlation between the stage of breast cancer and vascularization of lymph nodes was analyzed. The development of breast cancer was followed by rapid and significant increase in the number of blood vessel in axillary lymph nodes in general and in all their specific areas, especially in the paracortex and medullary substance. Strong or very strong positive correlation between the stage of tumor process and parameters of vascularization was observed in various areas of the axillary lymph nodes. No significant differences in node vascularization were observed between women without breast cancer and women with stage I breast cancer; the differences were found only starting from stage II. Considerable vascularization of lymph nodes can be a symptom of malignant tumor growth in the area of lymph collection. The duration of the tumor process and the size of the tumor can be indirectly evaluated by the increase in vessel number in lymph nodes.

Increased proteasome activity was revealed in blood serum of patients with stage T1N0M0 head and neck squamous cell carcinoma in comparison with patients with chronic diseases of the larynx and laryngopharynx. This opens prospects of using chymotrypsin-like activity measurement for differential diagnosis of squamous cell carcinoma, screening for high-risk groups, and evaluation of the degree of tumor differentiation

We studied the effect of pinealectomy on the parameters of 4-day infradian biorhythms in mature male Wistar rats. It was shown that pinealectomy did not change the period and phase of infradian biorhythm of total locomotor activity, serum concentrations corticosterone and testosterone, and mitotic index of esophageal epithelium. No significant changes in the number of nucleated cells in the spleen of pinealectomized animals in comparison with intact animals were found. The amplitude of biorhythms of locomotor activity and corticosterone and testosterone concentrations did not significantly change in pinealectomized animals. Despite the important role of the epiphysis in the regulation of circadian biorhythms in mammals, this organ is not the main synchronizer of infradian biorhythms of glucocorticoid and sex hormones in male Wistar rats.

Psychopharmacological effects of JNK inhibitor were studied using a mouse model of posthypoxic encephalopathy. The preparation exhibited a pronounced cerebroprotective effect manifested in normalization of orientation and exploratory behavior and conditioned responses in posthypoxic mice. These effects were accompanied by marked elevation of neural stem cell content in the paraventricular region of the brain.

Relationship between changes in the erythrocyte sedimentation rate in rats and concentration and charge of polyelectrolyte microcapsules was studied by the Panchenkov method. Positively charged microcapsules reduced erythrocyte sedimentation rate in a concentrationdependent manner. This effect was related to a decrease in the content of high-molecularweight proteins in the plasma due to their adsorption in positively charged microcapsules with polyacrylamide surface layer.

The effects of GK-1, a monomeric dipeptide mimetic of nerve growth factor (NGF) loop 4, on angiogenesis were studied in vitro and in vivo. Experiments on human umbilical vein endothelial cells HUVEC showed that the test compound did not affect tubulogenesis (initial stage of angiogenesis) and prevented realization of the angiogenic effect of NGF and its dimeric dipeptide mimetic GK-2. Experiments on rat hind limb ischemia model demonstrated that GK-1 (1 mg/kg/day intraperitoneally over 14 days) significantly reduced the density of the capillary network in ischemic tissue and increased the number and area of Zenker necrosis in comparison with the control. These data suggest that GK-1 exhibits a pronounced antiangiogenic activity.

The effect of allogenic combined transplantation of placental multipotent mesenchymal stromal and hemopoietic stem cells on regeneration of the myeloid tissue and spleen after acute blood loss was studied in laboratory mice. Combined transplantation of these cells did not change the content of cytogenetically modified cells in the bone marrow under normal conditions, but reduced their levels after acute blood loss. Combined transplantation of multipotent mesenchymal stromal and hemopoietic stem cells promoted activation of erythropoiesis and granulocytopoiesis. The major morphometric and cytological parameters of the white pulp of the spleen decreased, presumably due to immunosuppressive effect of multipotent mesenchymal stromal cells.

Activity of cathepsin D and phagocytosis of macrophages from vaginal lavage fluid, peritoneal exudation, and spleen were studied in mice of sensitive (DBA/2) and resistant (BALB/c) lines after intravaginal infection with type 2 herpes simplex virus and vaccination. Activity of cathepsin D and intensity of phagocytosis (irrespective of the macrophage source) and their ratio in BALB/c mice in early terms after infection were close to the control levels taken as a unit. In DBA/2 mice, these parameters and their balance were shifted and changes in cathepsin D activity depended on the time after challenge. Activities of cellular and extracellular cathepsin D increased sharply on day 1 postinfection under conditions of local virus interaction with the vaginal mucosa and activation of the pathological process. Later, after generalization of the infection, activity of cathepsin D decreased, while phagocytosis increased in all the studied macrophage populations. Vaccination corrected the cathepsin D/phagocytosis imbalance and created conditions for rapid elimination of the virus.

We studied association of single-nucleotide SCN5A (rs1805124), GJA5 (rs35594137), and KCNN3 (rs13376333) polymorphisms and sudden cardiac death. Humans died suddenly from cardiac causes (N=379) and unrelated sex- and age-matched control subjects were genotyped. No significant intergroup differences were found in the frequency of rs1805124 and rs13376333 genotypes and alleles. In women under 50 years, enhanced risk of sudden cardiac death was associated with rs35594137 GG genotype (OR=3.6; 95%CI=1.2-10.4; p=0.022), while in older women it was associated with rs35594137 AA genotype (OR=3.0; 95%CI=2.3-3.9; p=0.041). In women under 50 years, GA rs35594137 genotype was associated with a protective effect against sudden cardiac death (OR=0.3; 95%CI=0.1-0.8; p=0.036). Thus, GJA5 gene rs35594137 polymorphism is significantly associated with sudden cardiac death in the examined group.

Effects of bone marrow multipotent mesenchymal stromal cells and their secretory products released into the conditioned medium on microcirculatory bed in the broad ligament of the uterus were studied in Wistar rats with chronic genital inflammation. Opposite changes in the parameters of microcirculation and lymphatic drainage in the broad ligament of the uterus were observed after administration of cells and conditioned medium via different routes, which should be taken into account during the treatment of inflammatory and degenerative processes in the pelvic organs.

Individual features of the response of the hypothalamic—pituitary—adrenal axis (HPAA) to repeated moderate stress exposure (daily 2-h restraint stress for 10 days) was studied in young female rhesus monkeys with healthy normal behavior and combined group of female rhesus monkeys with abnormal depression-like and anxious behavior. No between-group differences in the response of ACTH and cortisol were found on day 1. On day 10, a rapid and less pronounced increase in ACTH secretion was observed in all animals in comparison with day 1. Analysis of between-group differences in HPAA response showed higher increase in ACTH level and lower increase in cortisol concentration in animals with depression-like and anxious behavior. These changes were similar to the previously described differences in the response of the adenohypophysis and adrenal cortex to acute restraint stress in old monkeys with similar behavior. Thus, individuals with depression-like and anxious behavior demonstrate impaired stress-induced reactivity of HPAA as early as in young age.

We studied the effects of single administration of a suspension of magnetomicelles based on carbon-coated iron nanoparticles on the structure of rat lungs within 40 days. Histological analysis revealed a complex of hemodynamic alterations in the lungs. Described changes persisted in the lung stroma from day 1 until day 40, but their intensity decreased by the end of the experiment. Using immunohistochemical Perls reaction we identified cells morphologically corresponding to alveolar and interstitial lung macrophages. The number of Perls⁺ cells decreased by day 40 of the experiment. Ultrastructural analysis showed endocytosis of modified iron nanoparticles and their accumulation in intracellular digestionorganelles (endo- and lysosomes) of mononuclear phagocyte system cells. Accumulation of magnetomicelles in the lungs was not associated with damage to pneumocytes, macrophages, and blood-air barrier.

We studied internalization of vector nanocarriers loaded with plasmid DNA into C6 glioma cells. For improving selectivity of plasmid delivery, the liposomes were conjugated with monoclonal antibodies to VEGF and its receptor VEGFR2. Flow cytofluorometry and laser scanning confocal microscopy showed more intensive (more than 2-fold) internalization and accumulation of antibody-vectorized liposomes in C6 glioma cells in comparison with the control (liposomes conjugated with non-specific antibodies and non-vectorized liposomes). Using quantitative analysis of fluorescent signal, we showed that cationic immunoliposomes significantly more effective delivered pCop-Green-N plasmid DNA and ensured effective transfection of C6 glioma cells.

Collagen I gels with protein concentrations of 1, 2, and 3.5 mg/ml were prepared and embedded in a porous polylactide scaffold to reduce their contraction. Concentration of the gel did not affect its degradation. Collagen gels promoted the formation of cell networks. The cells in the collagen gel with a concentration of 1 mg/ml embedded in polylactide scaffold had elongated spindle-like shape, in contrast to flattened cells in collagen gel of the same concentration not embedded in the scaffold. Stabilization of the collagen gel in the polylactide scaffold promoted active synthesis of laminin and fibronectin by cells as soon as on day 5 of culturing in comparison with that in free collagen substrate.

In 58 patients with coronary heart disease, the count of CD34⁺CD133⁺CD309⁺ endothelial progenitor cells in the blood was determined and the dynamics of the content of endothelial progenitor cells, angiogenic growth factors, and lipid parameters over 3 months of atorvastatin therapy was analyzed. Atorvastatin was administered in daily doses of 10 mg (26 patients) and 40 mg (32 patients). Control group comprised 15 healthy volunteers. In patients with coronary heart disease, the count of endothelial progenitor cells was lower by 4 times, the level of VEGF was higher by 52%, and the level of endostatin was lower by 13% than in healthy volunteers. Atorvastatin therapy significantly reduced the levels of VEGF (by 11%), C-reactive protein (by 26%), total cholesterol (by 30%), LDL cholesterol (by 35%), and triglycerides (by 18%); the levels of endostatin, MCP-1, and HDL cholesterol remained unchanged. The count of endothelial progenitor cells increased significantly by 72% irrespectively on the statin dose, but the changes were more pronounced in patients with lower initial endothelial progenitor cell counts and in patients with more drastic decrease in LDL cholesterol.

The initial stages of angiogenesis in rats after transcutaneous injection of autologous bone marrow multipotent mesenchymal stromal cells transfected with GFP gene and stained cell membranes in the projection of ligated femoral vein were studied by fluorescent light and confocal laser microscopy. Large clusters of brightly fluorescing elongated fibroblast-like cells were seen in the paravasal tissue and in the postoperative scar and signs of angiogenesis were noted as soon as in 4 days. The injected cells not only formed new vessels, but also integrated into vessels formed by host cells. Some injected cells were phagocytizied by macrophages and the latter started to fluoresce due to the presence of the membrane dye. These macrophages within the specified period appeared in the regional inguinal lymph nodes where they formed clusters in the lymphoid parenchyma of the cortical substance.

We studied the effect of nanostructured silicon dioxide particles on functional activity and ultrastructure of rat multipotent mesenchymal stem cells. A dose-dependent decrease in cell adhesion to plastics and their proliferative potential, accumulation of apoptotic cells, a decrease in the electron density of cell cytoplasm due to swelling and degenerative changes of organelles, and intracellular accumulation of nanostructured of silicon dioxide particles were observed.

We compared the production of 19 humoral factors in mixed cultures of mesenchymal stromal cells from Wharton’s jelly and allogenic peripheral blood lymphocytes. For evaluation of the specificity of immunosuppressive activity of mesenchymal stromal cells, comparative analysis of the production of these humoral factors in mixed cultures of lymphocytes and epithelial BxPC-3 cells was conducted. The production of soluble factors in both mono- and mixed cultures significantly correlated (p<0.05). The maximum production was found for proinflammatory chemokine IP-10 and IFN-γ and anti-inflammatory cytokine IL-10. The major difference of mesenchymal stromal cells from epithelial BxPC-3 cells was 7-fold higher production of IL-10, which can explain the immunosuppressive effect of mesenchymal stromal cells.