Vol 166, No 4 (2019)

We developed an approach for quantitative assay of injected vasopressin in urine samples by ELISA under conditions of physiological suppression of hormone secretion from the neurohypophysis into the blood. In experiments on unanesthetized rats, water load (5 ml/100 g body weight) almost completely blocked secretion of arginine-vasopressin. Injection of arginine-vasopressin in a dose of 0.1 nmol/100 g body weight after water load enhanced reabsorption of solute-free water and renal excretion of Na⁺, K⁺, and Mg²⁺ by 13.3, 5.5, and 5.0 times, respectively; urinary excretion of Ca²⁺ remained unchanged. It was found that urinary excretion of arginine-vasopressin directly correlated with reabsorption of solute-free water and renal sodium excretion.

We studied the influence of blockers of muscarinic M1, M2, and M3 receptors on the effect of acetylcholine in the myocardial tissue of caval veins in rats at the early stage of ontogeny. The experiments were performed on isolated preparations of the right superior vena cava working under their own rhythm. Action potentials were recorded using the standard microelectrode technique. Acetylcholine (1 μM) suppressed automatic activity in the superior vena cava myocardium. Preliminary perfusion of the preparation with non-selective blocker atropine (1 μM) completely abolished the effect of acetylcholine, treatment with M2 receptor blocker AQ-RA 741 (1 μM) led to partial suppression of the effect of acetylcholine. Blockers of M1 and M3 receptors pirenzepine (1 μM) and 4DAMP (0.1 μM) did not suppress the effect of acetylcholine. Thus, the effect of acetylcholine is predominantly realized via M2 receptors, but M3 receptors can also partially mediate its effect in the superior vena cava myocardium in rats at the early stages of ontogeny.

The development of arterial hypertension in male Wistar rats with fructose-induced metabolic syndrome (12.5% of fructose solution as the only drinking source for 10 weeks) along with impaired glucose tolerance and increased serum concentration of triglycerides and LPO products caused a decrease in the content of serum blood calcitonin gene-related peptide (CGRP). Low-frequency transcutaneous electrical nerve stimulation (1 mA, 2 Hz, 10 min daily for 2 weeks) performed in 8 weeks after the beginning of fructose treatment reduced systolic BP and serum concentration of triglycerides and LPO produces and improved glucose tolerance. After stimulation, CGRP content in rats maintained on fructose diet returned to normal values and the content of nitric oxide metabolites increased. We hypothesize that CGRP and nitric oxide are involved in mechanisms mediating the therapeutic effect of low-frequency transcutaneous electrical nerve stimulation on arterial hypertension developing in metabolic syndrome.

The effects of a natural complex of cytokines IL-1, IL-2, IL-6, TNF, MIF, and GTFβ on myocardial blood flow were studied under control conditions and during acute experimental aortal stenosis. Systemic administration of the cytokine complex under control conditions led to moderate impairment of the blood flow in the myocardium associated with plethora and perivascular edema. The number of functioning vessels in the myocardium significantly increased under these conditions, which reflected enhancement of the coronary blood flow. The comparison of the myocardial blood flow under conditions of acute heart overload alone and in combination with systemic administration of the cytokine complex revealed similar changes. In both cases, moderate plethora in all compartments of the vascular network, moderate perivascular edema, and moderate blood stasis in the myocardial capillaries were seen. The only difference the increase in the density of functioning capillaries that was significantly more pronounced after cytokine administration. These data indicate that the increase in the blood cytokine level induced dilatation of myocardial vessels and intensification of blood flow in normal and under conditions of acute hemodynamic heart overload. Against the background of pronounced vasodilatation, the dyscirculatory changes in the myocardium were moderate. It was assumed that the increase in the duration or frequency of hypercytokinemia episodes can induce more severe blood flow disturbances in the myocardium.

Pronounced analgesic activity of the innovative compound 4-(3,4-dibromthiophencarbonyl)-2,6,8,12-tetraacethyl-2,4,6,8,10,12hexaazatetracyclo[5,5,0,0^{3, 11},0^{5, 9}] dodecane (thiowurtzine) was observed in the thermal nociceptive hot plate test and in the acute visceral and somatic deep pain model (acetic acid writhing test). In these experimental models, naloxone-sensitive thiowurtzine-induced analgesia was revealed. The absence of tropism to peripheral opioid receptors in the acetic acid writhing test was demonstrated using naloxone methiodide. Course administration of low-toxic thiowurtzine in effective doses was not associated with ulcerogenic damage to the gastric mucosa in experimental animals.

In 24 h after combined administration of ligands of NOD2 (muramyl dipeptide) and TLR4 (LPS) receptors to CBA mice, a synergistic increase (by 10 times compared to the intact control) in cloning efficiency and content of multipotent stromal cells was observed in the bone marrow in comparison with the total effects of their individual administration (by 2.1 and 4.1 times, respectively). A similar effect was also observed in the peritoneal exudate. When ligands were administered simultaneously, the concentration of osteogenic multipotent stromal cells in the bone marrow decreased to a greater extent than in case of individual injections of the ligands, but did not drop below 7% of the control, which is apparently indicative of a decline threshold. In 3 h after simultaneous addition of the ligands in vitro to 12-day primary cultures of mouse bone marrow stromal cells, a synergistic increase in TNFα concentration was observed (32-fold increase from the level of intact control), while IL-10 concentration did not differ from the control, which is indicative of the proinflammatory nature of the process and the absence of immunosuppressive effect. These results suggest that activation of the stromal tissue depends on the intensity of innate immunity reactions.

The effects of ROS on functional properties (cytotoxic activity, antibody-producing activity, TNFα synthesis, and free cytosol calcium level), membrane structure (by expression of some surface markers), and apoptosis of lymphocytic cells were estimated in the peripheral blood of healthy volunteers. ¹O₂, \( {\mathrm{O}}_2^{\underset{\_}{\bullet }} \), OH^•, and H₂O₂ mostly suppressed cytotoxic activity of lymphocytes against Ehrlich ascites carcinoma cells and inhibited IgG synthesis and expression of receptors and surface markers (Fc receptors, CD3, CD19, and CD56). The exposure of lymphocytes to H₂O₂ (10^—6 M), ¹O₂, and OH^• was followed by an increase in the level of a secondary messenger, intracellular calcium, in comparison with non-exposed cells. The presence of exogenous calcium in the medium for lymphocyte suspending induced an increase in the number of cells at early and late stages of apoptosis 6 h after exposure to H₂O₂ and ¹O₂ in comparison with lymphocytes incubated in Ca²⁺-free medium.

We compared available methods for monitoring the integrity of in vitro models of barrier tissues and studied the possibility of using impedance spectroscopy to solve this problem. It was demonstrated (theoretically and experimentally) that TEER measurements are not sufficiently sensitive to detect small defects in the cell barrier that significantly affect its permeability. For obtaining reliable results, it is necessary to set a sufficiently high threshold TEER, which leads to the loss of many intact samples. At the same time, impedance spectroscopy has all advantages of the classical method of measuring TEER (it is rapid and non-invasive method), while its application in combination with the methods of machine learning allows reliable detection of defects in the cell barrier.

Production of cytokines and growth factors by cultured human umbilical cord tissue- and bone marrow-derived multipotent mesenchymal stromal cells was measured by multiplex analysis. In most cases, the concentrations of bioactive factors in the culture medium conditioned by umbilical cord-derived cells was ten- to hundred-times higher than in the medium conditioned by bone marrow-derived cells. These results suggest that both multipotent mesenchymal stromal cells from the umbilical cord and cell-free products can have more pronounced therapeutic effect in comparison with mesenchymal stromal cells obtained from “adult” sources.

We evaluated cytotoxic activity of peripheral blood NK cells towards trophoblast cells. NK cells either isolated or in the composition of mononuclear cell fraction, caused death of trophoblast cells. In women with physiological pregnancy, the cytotoxic effect of NK cells present in mononuclear cell fraction preincubated with IL-2 was lower than in nonpregnant women, who had never been pregnant previously, and in fertile women. Cytotoxic activity of isolated NK cells preincubated with IL-2 in fertile women was lower than in nonpregnant women, who had never been pregnant previously.

We performed a complex study of the dependence of immediate reaction of catalytic subunits in mitochondrial enzymes (NDUFV2, SDHA, Cyt b, COX1, and ATP5A) in rat cerebral cortex (the most hypoxia-sensitive tissue) on the severity and duration of hypoxia in vivo and the role of individual resistance of rats to oxygen deficiency in this process. Three types of responses to hypoxia were revealed. The immediate response of mitochondria to oxygen deficiency appeared after its drop by 30-33% relatively to normal atmosphere level. It manifested in up-regulation of NAD-dependent oxidation, i.e., activation of respiratory chain complex I. Further decrease in oxygen concentration by 50% reprogrammed the work of respiratory chain via activation of respiratory chain complex II in parallel with down-regulation of the electron transport function of the respiratory chain complex I. This response was optimal for the expression of adaptation genes and for the formation of immediate tolerance of rats to hypoxia. The greatest drop of oxygen concentration by 60-62% reversed the Krebs cycle promoting recovery of the electron transport function of respiratory chain complex I. Despite this, the energy efficiency of the respiratory chain and the potency to mobilize the rapid adaptation mechanisms degraded due to abnormalities in cytochrome segment of the respiratory chain.

The study focuses on the development of principally novel priority-oriented healthcare strategy — targeted therapy in regenerative medicine known as Strategy of Pharmacological Control over Intracellular Signal Transduction in Regeneration-Competent Cells. It implies selective action of promising drugs on specific key elements in the signaling cascade responsible for functional activity of various progenitor cells (including stem cells) and elements of tissue microenvironment. The results of pioneer studies are described that were aimed on revealing the peculiarities in signal transduction and the role of distinct signaling molecules (the potential targets) in the control of cell cycle and other functions of progenitor elements and regulatory cells of different types. The models of some pathological states were employed to demonstrate the possibility of effective implementation of the advanced pharmacotherapeutic concept. The developed theoretical and applied platform can be used to launch synthesis of principally novel preparations with regenerative activity.

Correlations between extracellular matrix components in mouse liver revealed the pathogenetically determined dynamic structure of these interrelations (some correlations appeared while others disappeared) at various terms of BCG-induced granulomatosis. The correlations between the studied parameters were strong during the first (postinfection days 3-10) and the second (postinfection days 60-90) periods but became moderate during the third period manifested by chronic inflammation (postinjection day 180). The greatest number of correlations was observed at the beginning of the stabilization period (postinjection day 60). At this period, the content of all structural components of hepatic proteoglycans closely correlated with that of hydroxyproline. These findings attested to systemic changes in metabolism of extracellular matrix components and interrelation between some structural units of proteoglycans in the liver and the components of extracellular matrix in the lungs. The common and typical organ-specific correlations in the liver and lungs suggest the existence of correlations between different organs during progression of BCG-induced granulomatosis.

The regulatory and adaptive status was determined in 202 healthy subjects by the parameters of the cardiorespiratory synchronism probe. We performed molecular-genetic analysis of polymorphic variants of the main gene of serotonin biosynthesis, tryptophan hydroxylase TPH1 (A218C polymorphism) and TPH2 (G703T polymorphism), and serotonin receptors (HTR2C and HTR2A genes). The association of the regulatory and adaptive status of a subject with the polymorphism of serotonergic mediator system genes was revealed.

We propose an approach that allows simultaneous determination of the levels of M and G isotypes of antibodies to the panel of glycans using microarrays. The level of IgG antibodies to 3’-O-su-Le^a glycan detects patients with colorectal cancer with a sensitivity of 69.77% and specificity of 62.75%. The percentage of correctly classified colorectal cancer patients with the use of a combination of two markers IgM antibodies to glycans 3’-sialyl-TF and 3’-O-su-Le^a is as high as 74.23%. The levels of IgM antibodies to 3’-O-su-Le^a glycan differ significantly in patients with and without regional metastases. The levels of some of antiglycan IgM or IgG antibodies differed significantly in patients with tumors of different location and differentiation.

We present prognostic risk models for structural and functional cardiovascular disorders in Caucasian adolescents (Russians) with essential hypertension aged 14-17 years (15.92±1.12) taking into account biochemical (catecholamines and cortisol) and genetic parameters (carriage of polymorphic variants of α2A-adrenergic receptor gene ARA2A (-1291С>G), norepinephrine transporter NET (-1287G>A), and D2-dopamine receptor DRD2 (-141C Ins/Del) and the data of psychological testing. Prognostically significant risk factors for increased specific peripheral vascular resistance and variability of daytime systolic BP were a combination of [CC+AA] genotypes of ADRA2A -1291C>G and NET -1287G>A polymorphisms, cortisol level, anxiety, and proneness to conflict. The combination of genotypes [CC+GG+II] of polymorphisms -1291C>G ADRA2A, -1287G>A NET and -141C Ins/Del DRD is prognostically significant, with the GG genotype NET being epistatic.

The abdominal type of cryptorchism was modeled on random-bred albino rats by replacing both testes from the scrotum into the abdominal cavity for 3 weeks; thereupon they were manipulated into the scrotum. In control rats, no additional surgery was performed. In experimental rats, the testicular tissue obtained from 1-2-day rat pups was transplanted under testicular tunica albuginea. Prior to orchiopexy, the weight of testes decreased by 62.5-64.1%. In 6 month after the surgery, it increased by 36.1% in the control group, whereas in experimental rats the weight of testes elevated by 123.2% and approximated the normal value. Histologically, the control group demonstrated persistent disturbance in spermatogenesis with emptiness of numerous seminiferous tubules where only Sertoli cells could be revealed and with pronounced dystrophic alterations in the spermatogenous epithelium of the partially preserved tubules where spermatogenesis was blocked at the spermatogonial level. In contrast, the transplantation region of the experimental testes exhibited formation of novel mature testicular tissue enclosed by a connective tissue capsule incorporating the seminiferous tubules with differentiated epithelium and with the clusters of Leydig cells in the stroma. In 6 month, spermatogenesis was observed in most seminiferous tubules of the host testicular tissue, which had spermatozoa in the lumens. To the moment of orchiopexy, the blood testosterone decreased by about 2.5-fold. In control group it remained diminished during entire observation period (up to 6 month), while in the experiment group its level normalized completely as early as in 2 month and remained even elevated to the end of observation period.

Immunohistochemistry of tissue factor, vimentin, and cytokeratin-8 was studied in the medical abortion material at gestation week 5-10 in 20 healthy women. No expression of tissue factor was found in vimentin-positive decidual cells of the parietal endometrium. By contrast, intensive immunostaining of the TF/FVIIα complex was detected in the marginal layer of Rohr fibrinoid and less intense staining was found in vimentin-positive decidual cells adjacent to the walls of modified spiral arteries in the utero-placental region under conditions of invasion of the interstitial and intravascular cytotrophoblast (marker: cytokeratin-8). In the villi, tissue factor was expressed only in the fibrinoid lumps at sites of the syncytiotrophoblast defects. We demonstrated the formation of local hemostasis system in the uteroplacental region at weeks 5-8. This system creates optimal conditions for external thrombogenesis aimed at realization of the cytotrophoblast invasion and subsequent flow of arterial blood between the villi at the end of the first trimester.

This paper presents the results of studying the effectiveness of the developed method for diagnosing the functional state of the dental system. The results indicate that the accuracy of the method is about 80%. The results of previous studies are reviewed and the prospects for diagnosis of the functional state of the dental system as a research and practical trend in dentistry are outlined. The physiological regularities of the dental system functioning and relationship between its functional status and the autonomic nervous and postural systems prompt a new trend of research, functional diagnosis in dentistry and interpretation of the findings of functional studies. The prospects of functional diagnosis in dentistry are discussed.

Mesenchymal stromal cells from rat adipose tissue were transduced with adeno-associated viral (AAV) vector encoding stem cell factor SCF that stimulates proliferation of cardiac c-kit⁺ cells and improved cardiac function and survival of animals after myocardial infarction. Extracellular vesicles isolated from the medium conditioned by mesenchymal stromal cells by ultracentrifugation were characterized by Western blotting, transmission electron microscopy, nanoparticle tracking analysis, immunostaining, and mass spectrometry analysis. Using proteomic analysis, we identified transgenic SCF in extracellular vesicles released by AAV-modified mesenchymal stromal cells and detected some proteins specific of extracellular vesicles secreted by transduced cells. Extracellular vesicles from AAV-transduced mesenchymal stromal cells could be used for delivery of transgenic proteins as they were readily endocytosed by both cardiosphere-derived cells and cardiac-progenitor cells.

Induction of direct cell death is one of the mechanisms of the antitumor effect of GD2-specific antibodies used for the therapy of high-risk neuroblastoma. The mechanisms of the cytotoxic signal triggered by antibody binding to GD2 ganglioside on the surface of the tumor cell remain insufficiently studied. Using inhibitor analysis we demonstrated that actin microfilaments are involved in the cell death induced by GD2-specific antibodies. Specifically, a strong antagonistic influence of cytochalasin D on the cytotoxic effect induced by GD2-specific antibodies was demonstrated in GD2+ tumor cell lines, which was expressed in at least 20% increase in cell survival and a significant decrease of the fraction of cells with fragmented DNA.

We studied the effect of mesenchymal stem cells and hemopoietic stem cells co-transplanted in different regimens (sequence and intervals between the injections within 0-48 h) on the rate of the bone marrow hemopoiesis recovery in 468 inbred mice with cytostatic aplasia caused by a single injection of cyclophosphamide in a dose of 500 mg/kg. The efficacy of stem cells was evaluated by animal survival and general physical condition, body weight dynamics, peripheral blood counts (leucocytes, platelets, and reticulocytes), total cellularity and the presence of hemopoietic stem cells in the bone marrow on day 9 after cyclophosphamide injection. The relative content of hemopoietic stem cells in the total myelokaryocyte pool was assessed by flow cytofluorometry using specific monoclonal antibodies to CD117 and CD184. The stimulating effect of co-transplantation of mesenchymal stem cells and hemopoietic stem cells on hemopoiesis was demonstrated by the indices of total cellularity and the relative content of CD117⁺ hemopoietic stem cells. The schemes with injection of mesenchymal stem cells 24-48 h prior to injection of hemopoietic stem cells were most effective.

We studied the influence of vascular prostheses made of polytetrafluoroethylene and polyethylene terephthalate on the proliferation, migration, and NO production by bone marrow mesenchymal stem cells, human endothelial progenitor cells, and EA.hy926 endothelial cells, colonization of the prosthesis surface by endothelial and mesenchymal cells was also analyzed. Synthetic prostheses have a negative effect on cell proliferation and migration, while surface treatment with proteins (fibronectin or gelatin) promotes colonization of the prostheses with cells.

The title of the article should read:

“Role of β Cell Precursors in the Regeneration of Insulin-Producing Pancreatic β Cells under the Influence of the Pegylated Form of Glucagon-Like Peptide 1”.