Vol 164, No 2 (2017)

In 1947 M. A. Bernstein proposed a hypothesis about “repetition without repetition” in biomechanics that was confirmed in psychophysiology by the Eskov—Zinchenko effect. This effect can be applied to all parameters (except the parameters of the neuromuscular system) of human body homeostasis. For instance, this instability can be demonstrated for repeated samples of cardiointerval parameters (and other homeostasis parameters) of the human cardiorespiratory system. Within the framework of the new theory of chaos and self-organization, a method is proposed for calculation of matrices of paired comparisons of cardiointerval samples for assessing the physiological status of the human body and changes in homeostasis. Statistical instability of cardiointerval samples and their statistical distribution functions f(x) for successive cardiointerval samples in one subject is proven.

We studied the effects of acute normobaric hypoxia on postnatal day 2 (model of preterm pregnancy) on reflex activity and behavior of juvenile male Wistar rats and the possibility of correction of behavioral deficit by administration of GABA derivative Salifen after hypoxia. It is shown, that perinatal hypoxia impaired righting reflex and forelimb grip strength and increased motor activity in juvenile male rats. Administration of Salifen for 14 days in a dose of 15 mg/kg improved reflex activity and behavior of rats, which indicates the prospect of further study of the therapeutic efficacy of this drug on models of neonatal encephalopathy.

Experimental studies demonstrated inhibition of glucose absorption in the jejunum in acute and chronic 1-h daily immobilization stress, with the maximum inhibition on day 7 of immobilization. These changes correlate with the development of oxidative stress in animals over the entire duration of the experiment, which manifested by an increase in the content of the primary and end LPO products and decrease of the total antioxidant activity of the blood. Correction of these shifts with the antioxidant drug inhibits accumulation of LPO products and increases antioxidant defense and glucose absorption rate. These data prove the important role of peroxidation in regulation of glucose absorption.

We studied the effect of Gly-His -Lys tripeptide administered intraperitoneally in doses of 5, 15, 50 and 150 μg/kg on pain-induced aggressive-defensive behavior. A foot-shock model of aggression in rats grouped in pairs in an electrified chamber was used. Analgesic and antiaggresiogenic effects of the peptide were demonstrated. It was found the L-lysine residue plays the key role in these effects, because they were observed under the influence of L-lysine administration in doses close to its equimolar content in the studied tripeptide.

Sulfated derivatives of xylan (isolated from Bétula pubéscens wood) with average molecular weight ~34 kDa, sulfur content of 11.3-17.5%, a degree of substitution of 0.74-1.64 are anticoagulants of direct type of action. Antithrombin and antifactor Xa activities in three tested xylan samples did not differ and reached 30.8-31.8 and 13.5-14.3 U/mg, respectively.

We studied the effects of a melatonin—aluminum oxide—polymethylsiloxane complex (complex M) on the expression of apoptosis regulators Bcl-2 and Bad in the liver of homozygous db/db BKS.Cg-Dock7^m+/+Lepr^db/J mice with obesity and type 2 diabetes. Complex M or placebo was administered daily through the gastric tube during weeks 8-16 of life. In mice with type 2 diabetes mellitus receiving placebo, enhanced immunohistochemical reactions for proapoptotic Bad protein and weak response for anti-apoptotic Bcl-2 protein were observed. Administration of complex M shifted the ratio of apoptosis regulators: the area of Bcl-2 expression significantly increased and against the background of reduced Bad expression area. These findings attest to antiapoptotic effect of complex M in the liver on the model of type 2 diabetes mellitus.

We compared the effects of GK-2 (dimeric dipeptide mimetic of nerve growth factor) and Mexidol (standard preparation for the therapy of stroke) on rat model of transient occlusion of the middle cerebral artery. GK-2 and Mexidol were administered intraperitoneally in the most active doses (1 and 100 mg/kg, respectively) 6 h after surgery and then once a day for 6 days. The preparations reduced the volume of cerebral infarction (by 60 and 30%, respectively). At the same time, GK-2 had a pronounced and statistically more reliable effect in a dose that is lower by two orders of magnitude. In addition, GK-2 significantly reduced the neurological deficit in the limb placement test, while Mexidol was ineffective in this test.

In untreated rectal cancer patients, the chymotrypsin-like activity of proteasomes in tumor tissue was 3-fold higher than that in conventionally normal tissue, which is explained by up-regulation of expression of immunoproteasomes and total pool of proteasomes. After neoadjuvant chemoradiation therapy, expressions of the total pool of proteasomes and immunoproteasomes in the tumor as well as the relative ratios of these indices to those in conventionally normal tissue were smaller by 1.4-3.3 times in comparison with the untreated patients. These changes were paralleled with pronounced (4.5-fold) down-regulation of proteasome activity in the tumor and a 3.7-fold decrease of activity ratio for the proteasomes in tumor and in conventionally normal tissue. The number of immunoproteasome subunits and the chymotrypsin-like activity of proteasomes can be viewed as potential markers to prognosticate effectiveness of neoadjuvant chemoradiation therapy in rectal cancer patients.

We compared changes in the redox status and intensity of oxidative modification of proteins in intact Jurkat tumor cells and cells cultured with buthionine sulfoximine, an inhibitor of the key enzyme of glutathione synthesis γ-glutamylcysteine synthetase. The glutathione system components play a role in modulation of the content of protein-bound glutathione, protein carbonyl derivatives, bityrosine, and oxidized tryptophan, and in dysregulation of apoptosis in Jurkat tumor cells. Inhibition of de novo synthesis of glutathione in Jurkat tumor cells was followed by accumulation of hydroxyl radical, a reduction in the level of protein-bound glutathione and oxidized tryptophan, and a rise in the concentration of protein carbonyl derivatives. These changes were accompanied by activation of programmed cell death.

A cascade of pathological changes in the intact hemisphere developed in rats 6 months after focal unilateral traumatic brain injury: neuronal degeneration, hyperexpression of α-synuclein, APP (β-amyloid peptide precursor) protein, and glutamine synthetase in cells other than astrocytes. The development of these changes in the contralateral hemisphere indicated the emergence of extensive delayed neurodegenerative processes in the brain after traumatic brain injury, which were characteristic of diseases associated with pathological aging.

The ultrastructure of nephrocytes of the proximal and distal convoluted tubules, podocytes, mesangial cells, and macrophages of the interstitial connective tissue was studied after single intravenous administration of magnetite nanoparticles modified with chitosan (magnetic nanospheres) or lipids (magnetic liposomes). Transmission electron microscopy showed ultrastructural features of absorption of magnetite nanoparticles. The shape, size, and number of vesicles containing nanoparticles in nephrocytes of convoluted tubules and macrophages after administration of the suspensions of magnetic nanospheres and magnetic liposomes were described.

Permeability of the blood—brain barrier for protein fractions 50-100 kDa (PF_50–100) of Cellex Daily preparation labeled with fluorescent tracer FITC and non-conjugated FITC were compared after intranasal administration of the preparations to healthy rats. Fluorimetrical analysis of the serum and cerebrospinal fluid samples showed that Cellex Daily PF_50–100-FITC administered intranasally penetrated into the blood and cerebrospinal fluid with maximum accumulation in 2 h after administration and persists in the circulation for 24 h probably due to binding with plasma proteins. The differences in the kinetic profile of PF_50–100-FITC and free FITC indirectly suggest that the major part of the preparation is not degraded within 24 h and FITC is probably not cleaved from the protein components of the preparation. In vivo fluorescence analysis showed significant fluorescent signal in the olfactory bulbs in 6 h after intranasal administration; hence, the preparation administered via this route can bypass the blood—brain barrier. Scanning laser confocal microscopy of rat brain sections confirmed penetration of the high-molecular weight protein fraction PF_50–100-FITC into CNS structures. The most pronounced accumulation of the labeled drug was observed in the olfactory bulb in 6 and 12 h after administration. In contrast to free FITC administered in the control group, significant accumulation of PF_50–100-FITC in the olfactory cortex and frontal cortex neurons with functionally active nuclei was observed in 6, 12 and 24 h after intranasal administration.

Course administration streptozotocin to male C57Bl/6 mice induces a complex of symptoms typical of type 1 diabetes mellitus: hyperglycemia and insulin deficiency, focal inflammatory infiltration of the pancreas, destructive changes in the Langerhans islets, damage to the insular apparatus (reduced number of PDX1⁺ cells and insulin expression by the secreting cells). Male reproductive disorder are serious complications of type 1 diabetes mellitus. In “diabetic” mice, interstitial edema with inflammatory infiltration and microvascular disorders in the testicular tissue are observed, the number of endothelial precursors (CD45^—/CD31⁺) and the total number and percentage of motile spermatozoa decreased, immature spermatogenic epithelium cells are desquamated of into the lumen of the tubules. Disturbances in the proliferation and differentiation of various spermatogonial stem cell populations (c-kit^—/CD90⁺, c-kit⁺/CD90⁺, and CD51^—/CD24⁺/CD52⁺) in diabetes can be explained by the inhibitory influence of inflammatory factors on testosterone-producing Leydig cells.

Recombinant human bone morphogenetic protein-2 with an additional s-tag domain (s-tag-BMP-2) synthesized in E. coli is characterized by higher solubility and activity than the protein without additional s-tag domain, which increases the yield during purification and simplifies protein introduction into the osteoplastic materials. The high osteoinductivity of the demineralized bone matrix with s-tag-BMP-2 was shown on the model of regeneration of cranial defects of a critical size in mice and on the model of implantation of porous titanium matrix into defects of femoral and tibial bones in rabbits.

Activity of hemantane, an amino adamantane derivative, exhibiting the properties of lowaffinity non-competitive NMDA receptor antagonist, was evaluated in experimental in vivo models of alcoholism. Hemantane had no effects on the formation and manifestation of behavioral sensitization to ethanol in DBA/2 mice. Under conditions of free choice between 10% ethanol and water, hemantane (20 mg/kg/day for 14 days, intraperitoneally) significantly reduced the daily ethanol intake in random-bred male rats with formed alcohol motivation (>4 g/kg of ethanol). During modelling of withdrawal syndrome, hemantane administered intraperitoneally in doses of 5-20 mg/kg dose-dependently attenuated alcohol-deprivation effect after acute withdrawal with no effects on protracted abstinence. It was found that hemantane suppressed alcohol drinking behavior in long-term ethanol experienced rats and attenuated alcohol-seeking behavior after acute withdrawal.

Association of TNF gene polymorphism -308G>A with the development of nonalcoholic steatohepatitis in the Russian population was revealed. Carriers of allele A of the TNF gene marker -308G>A have significantly higher risk of nonalcoholic steatohepatitis development: OR=1.69 (1.05; 2.71). Allele A carriage by this marker predicts an increase in the basal HDL level and a decrease in LDL and IL-10 levels in the blood of healthy subjects. Patients with nonalcoholic steatohepatitis, differing by the TNF gene -308G>A marker genotype, differ by the time course of the markers of hepatocellular damage (ALT, AST), activity of hepatocyte apoptosis (tissue polypeptide-specific antigen), and activation of specific humoral immunity (γ-globulin) in response to therapy with ursodeoxycholic acid in a dose of 10-15 mg/kg over 4-6 weeks. Carriers of allele A of the TNF gene polymorphic marker -308G>A are more sensitive to ursodeoxycholic acid therapy than carriers of GG genotype.

The anterior mediastinal lymph nodes were analyzed morphometrically in rats with chemically provoked breast cancer. Rats with untreated breast cancer and animals receiving chemotherapy demonstrated decreased volumes of paracortical region and lymphoid nodules with the germinal centers accompanied by extended medullary thymic substance. Resection of largest focus of breast tumor improved the filtration barrier potential of anterior mediastinal lymph nodes, up-regulated the proliferative activity of lymphoid cells in T-cell zones, and down-regulated proliferation of plasmatic cells.

Changes in the structure of the olfactory bulbs after long-term intranasal administration of pesticide rotenone, a classical inductor of parkinsonism, to rats were studied by the methods of immunomorphology. In rats intranasally receiving rotenone in a dose of 2.5 mg/kg every other day over 2 weeks, a decrease in the density of dopaminergic neurons and the area of astrocyte processes in the olfactory bulbs, activation of microglia in the glomerular layer, and enhanced α-synuclein phosphorylation and its accumulation in the bodies of mitral layer neurons were observed. The observed changes agree with the hypothesis on pathological α-synuclein transport via the olfactory route in Parkinson’s disease and confirm relevance of the rotenone model of Parkinson’s disease for studies of the pathological accumulation of α-synuclein.

The overwhelming majority of influenza vaccines are prepared with the use of chicken embryo allantoic fluid. The presence of ovalbumin (this protein constitutes >60% total protein in the allantoic fluid) in the vaccine can lead to severe allergy. Hence, effective reduction of ovalbumin content is of crucial importance for vaccine production. We compared two methods of purification and concentration of influenza virus: zonal gradient ultracentrifugation and combined ultrafiltration/diafiltration and exclusion chromatography protocol, used for fabrication of seasonal vaccines. Combined chromatography is comparable with zonal centrifugation protocol by the results of ovalbumin removal (to meet standard requirements).

We studied in vivo modifying effect of autotransfusion of human bone marrow mesenchymal stromal cells on ROS generation and production of cytokines (TNFα,TNFβ, IL-1α, IL-10, IFNγ, and GM-CSF) and PGE₂ by mononuclear cells of patients (N=21) with chronic heart failure. These parameters were evaluated prior to (control) and after (immediately and on day 14) intravenous administration of stromal cells in doses of 100-200×10⁶. Immediately after autotransfusion, significant increase of in vitro zymosan-induced chemiluminescence of blood mononuclear cells from 10 patients was observed. At later terms after autotransfusion (day 14), inhibition of chemiluminescent activity of blood mononuclear cells was revealed in 50% patients. We discuss possible mechanisms of involvement of transplanted autologous bone marrow mesenchymal stromal cells in reprogramming of blood mononuclear phagocytes from the pro- to anti-inflammatory phenotype under conditions of their in vivo interaction manifesting in transition from activation to inhibition of ROS-producing activity of macrophages and significant suppression of in vitro LPS-induced production of TNFα and GM-CSF by blood mononuclears against the background of significantly elevated TNFβ, IL-10, and IL-1α concentrations.

We studied structural and functional properties of human platelets in the presence of nanosilver particles. Incubation with 0.05-5 μM silver nanoparticles suppressed platelet adhesion in a dose-dependent manner without affecting internal platelet structure; during adhesion, some granules were not exocytized. Spontaneous platelet activation was observed at nanoparticle concentrations 15-100 μM. Addition of 1-5 μM nanosilver to cells undergoing adhesion blocked massive platelet degranulation, but did not prevent the formation of lamellopodia. The maximum number of preserved granules in platelets was revealed in the presence of 2.5-5 μM silver nanoparticles: 50% after platelet preincubation with silver nanoparticles and 75-77% after stabilization of adherent platelets with silver nanoparticles.

Huntington’s disease is a hereditary neurodegenerative disease that primarily affects striatal neurons. Recent studies demonstrated abnormalities in calcium regulation in striatal neurons in Huntington’s disease, which leads to elimination of synaptic connections between cortical and striatal neurons. In the present study, we focused on the neuroprotective properties of σ1-receptor, because one of its main functions is associated with modulation of calcium homeostasis in cells. The application of selective σ1-receptor agonists to the corticostriatal cell culture restores synaptic connections between the cortical and striatal neurons. Based on the obtained data, we assume that σ1-receptor is a promising target for the development of drugs for the therapy of Huntington’s disease.

The effectiveness of autologous cell product in the therapy of skin burn wounds was studied in C57B1/6 male mice against the background of streptozotocin-induced diabetes mellitus. In animals with and without modeled diabetes mellitus, significant decrease in skin defect area was observed after single administration of the cell product (bone marrow multipotent mesenchymal stromal cells, fibroblasts or media conditioned by these cells).

We performed comparative analysis of the morphology of chondrocytes in normal cartilage, after their isolation from the tissue, and at different stages of culturing; structural dynamics of cells during culturing was also studied. Significant morphological differences in chondrocytes at the specified stages of their preparation to in vivo use were revealed. Pronounced structural changes (blebbing and cytoplasm swelling) were found in chondrocytes before their implantation, which can affect the formation of cartilage regenerate. The study was performed using light microscopy methods including time-lapse recording of the cell cultures with differential interference Nomarski contrasting combined with transmission electron microscopy.