Vol 24, No 2 (2021)

Cover Page

Full Issue


Role of mesenchymal stromal/stem cells in regulation of hemotoration in 3D in vitro culture

Ivanov P.A., Yurova K.A., Khaziakhmatova O.G., Shupletsova V.V., Malaschenko V.V., Shunkin E.O., Norkin I.K., Khlusov I.A., Litvinova L.S.


The article is devoted to studying the role of mesenchymal stromal cells in formation of microenvironment for hematopoietic stem cells under the conditions mimicking physiological bone remodeling in presence of artificial three-dimensional matrices (Ra = 2-3 μm). The study was carried out using experimental samples of artificial implants obtained in electrolyte from hydroxyapatite nanoparticles (HAP) produced at the Institute of Strength Physics and Materials Science (Siberian Branch of the Russian Academy of Sciences). The work included cultural and instrumental research techniques. Phenotypic profile of cells was assessed by flow cytometry. Determination of cytokine/chemokine levels from cell culture supernatants was assessed by flow fluorimetry. Detection of hematopoietic cells in the vision fields, as well as areas of extracellular matrix mineralization was carried out by means of cytomorphometry.

It was revealed that the 3D matrices with a calcium phosphate coating initiate the in vitro formation of specific microenvironment of MSCs, resulting in the increased numbers of HSCs with the CD45+CD34+ phenotype (at 14 days), an increased number of cells with hematopoietic morphology and evolving foci of extracellular matrix mineralization of the (at 21 days). Changed numbers of hematopoietic cells per vision field occurred, mainly, due to indirect effect of hematopoietic factors (SCF and G-CSF), along with decrease of proapoptotic factor TRAIL. It was also found that MSCs reduce the level of proinflammatory cytokines (IFNγ, TNFα, IP-10, IL-2, IL-6) in culture medium in the presence of artificial 3D calcium-phosphate-coated matrices. The revealed features of MSC functioning under the conditions simulating physiological bone remodeling, upon co-cultures with three-dimensional matrices (Ra = 2-3 μm), have shown a significant effect of MSCs upon regulation of HSCs by local microenvironment, through distinct modulating effects of cytokines, chemokines, and growth factors that provide intercellular interactions. Development of extracellular matrix mineralization areas during MSC cultivation in the presence of 3D matrices imitating mineral substance of bone tissue also indicates the formation of osteoblastic niches under the in vitro cultivation conditions.

The results obtained are important in order to assess functions of hematopoietic niches and the role of MSCs in their development and maintenance of the microenvironment.

The results obtained may find practical application in development of new classes of medical devices able to provide effective osseointegration.

Russian Journal of Immunology. 2021;24(2):153-160
pages 153-160 views

Role of MP-4 myelopeptide in regulation of antibody production and functional activity of macrophages under stress conditions

Gavrilova T.V., Oralova D.A., Gein O.N., Chereshneva M.V.


Myelopeptides are endogenous low-molecular weight peptides with immunoregulatory activity. Each of these myelopeptides has an individual sequence and plays a role in correction of immune system disorders. In our earlier studies, we have shown immunomodulatory effect of MP-3, MP-5, MP-6 myelopeptides upon functioning of peritoneal macrophages under the conditions of immobilization and cold stresses. The aim of this work was to investigate the effect of MP-4 myelopeptide upon antibody formation, production of reactive oxygen species, and absorptive activity of peritoneal macrophages in mice under the immobilization stress. Materials and methods. The in vivo experiments were carried out in 24 male Swiss mice weighing 17-22 g. Two-or six-hour immobilization stress was produced in the animals. MP-4 was injected intraperitoneally 30 min before the onset of stress, at a dose of 40 μg/kg. The animals were divided into 4 groups: 1, control; 2, stress; 3, stress + MP-4; 4, MP-4 injection. On day +5, cellularity and the number of antibody-forming cells (AOC) in spleen were assessed by the method of local hemolysis in an agarose gel according to Jerne. Production of reactive oxygen species by peritoneal cells was assessed using the luminol-dependent chemiluminescence (LZHL) reaction. The absorption capacity of peritoneal macrophages was assessed using a standard method using a BDFACSCalibur laser flow cytometer. Labeled St. aureus at a final concentration of 108 cells/ml were used as phagocytosis targets. Statistical analysis was performed using the unpaired Student's t-test. Results. It was found that two-hour immobilization stress did not affect the absorption activity of peritoneal leukocytes, whereas six-hour stress suppressed phagocytosis. Injection of MP-4 in stress-exposed animals did not lead to changes in phagocytic activity of peritoneal macrophages. MP-4 did not correct the stress-induced decrease in the number of antibody-forming cells from the spleen during six hours of immobilization, nor stress-induced increase in absolute amounts of AOC under the two-hour immobilization stress. In the group of animals receiving MP-4 and immobilized for two-hours, an increase in the number of nucleated cells was increased, as compared with the control group. Thus, in contrast to the previously studied myelopeptides, MP-4 did not show pronounced immunomodulatory effects upon these parameters.

Russian Journal of Immunology. 2021;24(2):161-166
pages 161-166 views

Experimental evaluation of cytoprotective effect of probiotic metabolites and selection criteria of Bacillus Subtilis strains for development of new immunotropic biohepatoprotector

Zabokritskiy N.A.


The present work concerns a study of cytoprotective effect of biologically active substances (BAS) produced by probiotic microorganisms from the Bacillus genus. These substances were obtained from a sterile fugate of probiotic Bacillus subtilis VKPM B-3679 culture (a biosporicin-producing strain) and added to the culture of isolated hepatocytes, when modeling their in vitro toxic damage. Sterile buffered culture of Bacillus subtilis B-3679 was prepared by sterilizing filtration of the culture supernatant of this strain. The work was carried out using a passageable L-41 cell culture strain, which allows to assess toxicity of various substrates upon cell cultures and protection against toxic effects at different concentrations. At the first stage of study, the maximal non-toxic dose of sterile fugate and the minimal toxic dose of CCl4 established for the cultured L-41 cells. At the next stage, cytoprotective effect of BAS originating from the of B. subtilis B-3679 strain fugate, was studied and tested with L-41 cell line. Cytoprotective and regenerative effects of BAS containing in the fugate of B. subtilis B-3679 strain were demonstrated in the model of toxic damage using the culture of isolated hepatocytes. In preclinical studies, to assess the toxicity and safety of the experimental sample of a new biohepatoprotector for experimental animals, we have found that the complex of biologically active substances (metabolites) is non-toxic and safe for laboratory animals when administered intragastrically and intraperitoneally, and it does not cause any pathological changes in their internal organs and tissues. The basis of the new biohepatoprotection effect, which may provide a multifunctional action, allowing to effectively restore depressed liver functions, with simultaneous normalization of immunological parameters, is its active biocomponent, i.e., metabolites of probiotic spore-forming bacteria, which, when if brought to the body, produce a complex of biologically active metabolites (antibiotics, proteolytic, amylolytic, and other enzymes, immunoglobulins, as well as interleukins, vitamins, proteins, amino acids, and others bioactive substances). Due to experimentally established cytoprotective effect of the complex of BAS, the components of fugate culture of B. subtilis strain B-3679, will allow us to develop new promising medical immunobiological drugs that may provide a protective effect on human organs and tissues. As a result it was found that BAS, as components of fugate from B. subtilis B-3679 strain show both a pronounced cytoprotective effect, and a positive action upon regenerative abilities of liver cells, which is a significant factor for the future use of this strain as a biocomponent of a new immunotropic biohepatoprotector.

Russian Journal of Immunology. 2021;24(2):167-174
pages 167-174 views

Influence of glycodelin on morphofunctional state of secondary immune organs in experimental model of allogeneic transplantation

Loginova N.P., Troynich Y.N., Charushina Y.A., Zamorina S.A.


Glycodelin (PP14, PAEP, alpha-2-microglobulin, dimeric glycoprotein with a molecular weight of 42 to 56 kDa) is a marker of reproductive tissue receptivity. Immunoregulatory potential of glycodelin allows us to consider it among key factors that promote maternal immune tolerance to the developing embryo. In general, glycodelin has prospects for use in biomedicine as a biopharmaceutical to treat post-transplant complications. This work aimed to study the effect of glycodelin on morpho-functional state of the secondary organs of immune response in the course of in vivo experiment during allogeneic transplantation of bone marrow cell suspension to Wistar rats. The experiment was performed in white male Wistar rats aged 2-3 months (body mass, 250 g). The animals were kept at the Perm State National Research University animal clinic under GOST 33216-2014 “Rules for working with laboratory rodents and rabbits”. The animals were sacrificed on the 3rd, 7th, 14th, and 21st days according to international rules for working with experimental animals. In the control group, animals (n = 6) were injected once with a suspension of camptothecin-treated bone marrow cells; in experimental group, the animals (n = 12) were injected with a bone marrow cells in combination with glycodelin administered 4 times.

We used recombinant glycodelin (MyBioSource, Germany), which was administered to the animals at a concentration corresponding to pregnancy state (0.75 μg/mL). Intraperitoneal injection of bone marrow cells simultaneously with glycodelin over the terms of experiment (21 days) showed that allogeneic transplantation caused hyperplasia of spleen and lymph nodes. In the functional zones of organs, proliferation and differentiation of immune cells developed. Injection of glycodelin at the early stages of experiment (3-7 days) caused an increase in proliferative processes in the organs of both T- and B-dependent immunity. Moreover, there were no signs of inflammation and apoptotic cell death in the organs. Since the 14th day of the experiment, eosinophilic infiltration of the organs was evident, being an indirect positive sign of response to the transplanted cells. By the end of study, differentiation processes dominated over proliferation in the organs. Thus, glycodelin stabilizes proliferative processes and promotes emergence of a new cell generation, thus supporting host response to the transplant. Apparently, glycodelin can participate in development of adaptive mechanism in secondary organs of immune system.

Russian Journal of Immunology. 2021;24(2):175-180
pages 175-180 views

Effect of melatonin include into original dermal film upon adaptive immunity in experimental thermal trauma

Osikov M.V., Simonyan E.V., Ageeva A.A., Nikushkina K.V.


Adaptive immunity changes in thermal trauma (TT) increase the risk of infectious complications and limit repair of the lesion. Hence, search and preclinical testing of effective and safe means to locally manage TT, containing bioregulators, is an urgent task of modern medicine. Dermal films (DF) are an innovative and popular variant of wound coatings for small-area burns, and pleiotropic properties of melatonin (MT) suggest its effectiveness in TT. The aim of the work is to investigate the effect of MT, as a component of original DF, upon the indexes of adaptive immunity in experimental TT. The experiment was performed on 115 male Wistar rats. Grade IIIA TT with area of 3.5% were produced by contact with boiling water for 12 s. DF with an area of 12 cm2 based on sodium carboxymethylcellulose contatning MT (5 mg/g) was applied daily for 5 days. Similar DF matrix, but without MT, was used in the control group, Amounts of CD3+ and CD45RA+ cells in blood, and lymphocyte subpolulations with early and late signs of apoptosis and partially necrotic cells were evaluated with flow cytofluorometer, as well as IgG and IgM concentrations were measured in blood serum using rat test systems. With TT, the amount of CD3+ in the blood decreases on days +5 and +10, CD45RA+, on days +5, +10 and +20, and the concentration of IgG in the serum, on days +5 and +10 of observation. On days +5 and +10 after TT, a relationship was established between CD3+ and the number of lymphocytes with signs of early apoptosis (R = -0.47; p < 0.05; R = -0.51; p < 0.05, respectively), and signs of late apoptosis and necrosis (R = -0.64; p < 0.05; R = -0.42; p < 0.05, respectively), between CD45RA+ and the number of lymphocytes with signs of early apoptosis (R = -0.47; p < 0.05; R = -0.49; p < 0.05, respectively), and signs of late apoptosis and necrosis (R = -0.57; p < 0.05; R = -0.49; p < 0.05, respectively). Usage of MT in DF composition leads to increase in blood CD3+ on the 5th and 20th days, CD45RA+, on the 5th day, and an increase in serum IgG concentration was observed on the 5th and 10th days following TT. Restriction of necrotic and apoptotic death of blood lymphocyte may be among the mechanisms of the immunotropic effect produced by MT which is, probably, due to its local antioxidant and anti-inflammatory action in the TT area.

Russian Journal of Immunology. 2021;24(2):181-188
pages 181-188 views

Epiplexus phagocytes of nervous tissue in experimental brain contusion

Plekhova N.G., Zinoviev S.V., Prosekova E.V., Radkov I.V.


Neuroinflammation is known to participate in pathogenesis of intracranial brain injury (TBI), e.g., brain contusion or concussion. In view of high overall prevalence of these conditions, there is a need for nosological verification of the mild- and moderate-severity neuroinflammation. Our research in immune regulation of blood flow in TBI, including a role of Kolmer cells in pathogenesis of neuroinflammation, is now at the stage of collecting research data and requires adequate experimental study. Purpose of our study was as follows: to assess the state of Kolmer cells in vascular plexus of brain ventricles in experimental model of mild traumatic brain contusion (mTBC). mTBC was reproduced in male Wistar rats using a model of a falling load weighing 200 g. Immunohistochemical study was performed in order to assess CD45 receptor expression on the brain cells. During acute period after mTBC, we observed constriction of blood vessels and pericellular edema of the brain tissues. Expression of CD45 cytodifferentiation receptors markers characteristic of the hematopoietic cell pools was found in parenchymal areas of neocortex and on the surface of choroid plexuses in brain ventricles. These data suggest participation of epiplexus and parenchymal macrophages in the pronounced pericellular edema of the brain. On the 8th day of observation, the spasm of the blood vessels persists, along with significantly weaker pericellular edema. In all the brain sections, leukocyte infiltration of tissues was not seen, and there was no expression of CD45 receptors, whereas increased number and size of nucleoli was found in the neurons. The results of our study confirm the role of cerebral vasospasm as a severe complication of neuroinflammation developing after mTBI. Acute inflammation is characterized by a series of vascular changes, manifesting by development of vasospasm, arterial, venous hyperemia and stasis. Venous hyperemia is characterized by further vasodilation, tissue plethora, the phenomenon of the marginal leukocyte stasis and their emigration, along with increased exudation processes. Innervation from subcortical neurons or local cortical interneurons to parenchymal arterioles and cortical microvessels provides minimal contact and predominantly targets the surrounding astrocytes and other cells. During acute period of mTBC, the inflammatory process is confirmed by the presence in parenchyma and on the surface of ependymal Kolmer cells and by expression of CD45 receptors. This finding points to inflammatory reasons for altered tone of pial blood vessels, capillaries of neocortex and ependymal areas, and changed depth of subarachnoid space. Response of astrocytes to the brain trauma could be another factor of neuroinflammation.

Russian Journal of Immunology. 2021;24(2):189-194
pages 189-194 views

Reaction to damage of connective tissue in immunoprivileged organ (testis)

Khramtsova Y.S., Tyumentseva N.V., Artashyan O.S., Yushkov B.G.


Microenvironment of sperm and its precursors includes various immune cell populations. This indicates not only their importance for immune privileged state within testes, but it concerns a regulatory role of these structures in performance of the most important physiological functions. Despite sufficient knowledge on the immune privileged state in the organ, the regulatory function are scarcely studied, and existing literature virtually does not cover the issues of local spermatogenesis regulation by various components of testicular microenvironment in the course of their regeneration. Purpose of the present study was to define the reactions of connective tissue in rat testis following traumatic lesion. Materials and methods: the study was carried out in mature male Wistar rats. Experimental animals were divided into 2 groups: intact animals and animals with blunt trauma to the left testicle. The animals were removed from the experiment on the 7th and 30th days. Blunt trauma was simulated by squeezing the organ with forceps with a force of 15 N for 3 seconds. For histological examination, the testes were excised, preparations were made by the standard scheme, stained with hematoxylin/ eosin, toluidine blue (to identify mast cells), and according to Van Gieson (to detect collagen fibers). Distinct components of connective tissue and spermatogenesis were evaluated in testicular preparations. Quantitative indexes were calculated using the ImageJ program. Total testosterone levels in the blood were determined by chemiluminescence technique. Statistical evaluation was performed with Statistica 8.0 software. Comparison of groups was performed using Mann-Whitney test. We have found that restoration of spermatogenesis in the damaged testis did not occur within 30 days after the injury. While the reaction of connective tissue was noted in the both testes, it was more pronounced in the damaged organ, and manifests as changes in testicular microvasculature, stimulation of fibroblastic response, multidirectional effects of mast cells and Leydig cells, depending on the duration of exposure. Changes in various components of microenvironment in the damaged testis led to similar changes in the intact organ. The mechanism of this change is usually associated with effect of antisperm antibodies and development of autoimmune processes, but another possible mechanism for impairment of spermatogenesis in the second paired intact organ may include effects of connective tissue microenvironment upon the spermatogenic epithelial cells.

Russian Journal of Immunology. 2021;24(2):195-202
pages 195-202 views

Influence of different kinds of stress on apoptosis of CD4+/CD8+ lymphocytes upon in vivo blockage of opiate receptors

Sharavieva I.L., Gein S.V.


The endogenous opioid system is involved in regulation of many body functions, including immune system regulation. Endogenous opioid peptides are released under stress and communicate with opiate receptors, which are found on most immune cells. Apoptosis is one of the most important instruments in the regulation of the immune system. The aim of this study was to evaluate effects of various types of stress upon apoptosis of CD4+/ CD8+ lymphocytes under conditions of opiate receptor blockade. The objects of this study were white outbred male mice, been kept in laboratory vivarium. The following types of stress were studied: immobilization (6 h in the supine position), rotational (60 min: 10 min rotation at 78 rpm, 5 min rest), two types of acute cold (10 min or 60 min at -20 °С), chronic cold (-4 °С for 4 hours daily, for 7 days). Opiate receptors have been blocked by subcutaneous administration of naloxone hydrochloride at a dose of 0.2 mg/kg 20 minutes before stress, and again 3 hours later, with a stress duration of over 3 hours. Splenocytes were stained with PE-labeled monoclonal antibodies against murine CD4, and antibodies against mouse CD8 (BioLegent, USA); after incubation and washing, they were stained with reagents for determining apoptosis. V-FITC / 7-AAD annexin kit (Beckman Coulter, USA) according to the manufacturer's instructions. Lymphocyte apoptosis was recorded with a CytoFLEX S cytometer (Beckman Coulter, USA). Results. It was found that immobilization stress (lasting 6 h) and acute cold stress (60 min -20 °С), regardless of naloxone administration, enhance apoptosis of mouse CD8+ lymphocytes. Chronic cold stress (-4 °С 4 hours / 7 days) led to a decrease in apoptotic spleen lymphocyte numbers only during the blockade of opiate receptors. The effects of rotational stress and short-term acute cold stress (-20 °С, 10 min) upon apoptosis were not registered. Thus, two types of stress (immobilization and acute cold) caused intensification of CD8+ lymphocyte apoptosis; endogenous opioid system seems not to participate in regulation of these processes. Under conditions of chronic cold stress with naloxone injections, we have noted a decreased Annexin binding by spleen lymphocytes in mice.

Russian Journal of Immunology. 2021;24(2):203-208
pages 203-208 views

Autoimmune profile of rat blood in experimental ulcerative colitis

Davydova E.V., Osikov M.V., Bakeeva A.E., Kaigorodtseva N.V.


Worldwide incidence of digestive system disorders doubles each decade, thus representing a significant medical and social problem. Despite lacking knowledge in pathogenesis of inflammatory bowel diseases, it is clear that serum cytokine imbalance, and lesions in the walls of gastrointestinal tract are observed in experimental colitis. Pathogenesis of UC remains controversial due to a large set of etiological factors that initiate activation of cellular and humoral mechanisms of the immune response upon development of inflammatory changes in the large intestine. At the same time, cytokine secretion and expression have not been studied in details. The aim of the work was to study the cytokine profile of blood in rats using the experimental oxazolone model of ulcerative colitis. The work was performed in 40 white Wistar rats; ulcerative colitis was induced by rectal administration of a 3% alcohol solution of oxazolone. For anesthesia, Zoletil-100 (INN: tiletamine hydrochloride, VirbacSanteAnimale; France) was used at a dose of 20 mg/kg. The studies were carried out on the 2nd, 4th and 6th days. Serum concentration of IL-6, IL-8, IL-17 and IL-23 was determined by means of automatic ELISA analyzer “Personal LAB” using a specific test system for rats. For immunohistochemistry of Treg cells, we used anti-FoxP3 antibody (Arigo Biolaboratories, Тайвань) followed by immunhistostaining in VENTANA BenchMark XT (USA). Statistical evaluation was performed by non-parametric Mann-Whitney and Wald-Wolfowitz criteria. The difference was considered significant at р ≤ 0.05. In rats with experimental colitis, an increase of proinflammatory IL-17 which acts by attraction of neutrophils and other cells of innate immunity, supporting chronic inflammation and autoimmune reactions. We have found an increase of serum IL-23 concentration in rats with experimental ulcerative colitis on days 2, 4 and 6 of the experiment. This cytokine induces and maintains the inflammatory process in the wall of the large intestine. Significant decrease of FoxP3+Т-lymphocytes was revealed in colonic tissues, thus suggesting appropriate local autoimmune disorders.

Russian Journal of Immunology. 2021;24(2):209-214
pages 209-214 views

Effect of second-hand smoke on the immunological parameters of wistar rats and their offspring

Kuzmicheva N.A., Mikhailova I.V., Chaynikova I.N., Pushkareva L.A., Filippova Y.V., Smolyagin A.I.


The literature covers multiple negative effects of active and passive tobacco smoking upon the human body and experimental animals. Much lesser attention is given to the features of the immune system in offspring from the passively smoking experimental animals. In previous works, we studied physiological and immunological parameters in the infant rats born from passively smoking rats. The present work expands the scope of this direction and is devoted to assessment of immunological parameters in passively smoking pregnant rats and their offspring. Quantitative determination of immunological parameters was carried out in pregnant Wistar rats exposed to passive tobacco smoking and in 65 pup rats from smoke-exposed and nonexposed animals. The experimental rats were exposed to tobacco smoke for 8 hours from the 1st to the 20th day of pregnancy. In all animals, the body weight, thymus and spleen, the number of white blood cells, thymocytes, splenocytes, and myelocaryocytes were determined. On the 20th day of pregnancy, the smoking rats showed a significant decrease in the weight of animals, the number of thymocytes and myelocaryocytes. A decreased body mass of animals and their thymus glands, lower number of thymocytes were registered in pup rats born from smoking females as compared with control animals. These changes in immunological parameters may be based on a number of possible reasons. On the one hand, tobacco smoke promotes activation of free-radical processes with a simultaneously decreased efficiency of antioxidant mechanisms. This activation may result from exposure to the components of tobacco smoke, in particular, chromium, as a metal of variable valence, as well as benzene, which is transformed by the free-radical oxidation mechanisms. On the other hand, some tobacco smoke components may promote apoptosis which plays a significant role in the cellular exhaustion of immunogenesis system. Moreover, a decreased number of thymocytes may be caused by their migration from cortical areas to the medulla, followed by release into bloodstream. Thus, the results obtained indicate that the detected changes in these parameters are more pronounced in pregnant rats who were exposed to tobacco smoke and their offspring, and expand the knowledge of changes in the immune system in the offspring of passively smoking animals. The presented data may present a basis for development of drugs that reduce toxic effect of ecotoxicants, including passive tobacco smoking.

Russian Journal of Immunology. 2021;24(2):215-220
pages 215-220 views

Comparative analysis of bactericidal properties of synthetic peptides from the active center of GM-CSF - ZP2 against different gram-negative bacteria

Gritsenko V.A., Tyapaeva Y.V., Dobrynina M.A., Zurochka A.V.


Objective of this study was to carry out comparative analysis of bactericidal activity of synthetic peptide ZP2 (SP ZP2) against museum strains and clinical isolates of Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii. Materials and methods. We used museum strains of E. coli (ATCC 25922) and P. aeruginosa ATCC 27853, as well as 104 clinical isolates of Enterobacteriaceae, including E. coli (n = 22) and K. pneumoniae (n = 82), and 98 clinical isolates of non-fermenting Gramnegative bacteria, including P. aeruginosa (n = 43) and A. baumannii (n = 55), isolated from patients with various gynecological and surgical infections. Bactericidal activity of SP ZP2 (final concentration 10 μg/ml) against the microorganisms was assessed by difference in optical density (OD) for experimental and control broth cultures after 20 min of contact of bacterial suspensions (5 x 108 CFU/ml) with SP ZP2 (in control — with distilled water), adding meat-peptone broth and 4-hour incubation at 37 °C. The effect of SP ZP2 was expressed by the Bactericidal Activity Index (BAI, %). Results. Using the in vitro assays, we have found that SP ZP2 had a pronounced bactericidal effect on both the reference strains of E. coli and P. aeruginosa, and majority (95.5-98.2%) of the studied clinical isolates of Gram-negative bacteria, regardless of their species. With regard of the average BAI values, the tested bacterial species may be ranked by increasing their sensitivity to the synthetic ZP2 peptide as follows: P. aeruginosa (74.0±2.3%) — E. coli (77.6±3.5%) — K. pneumoniae (82.8±1.6%) — A. baumannii (84.3±1.7%). In addition, significant intraspecific variability of clinical isolates of Gram-negative bacteria was revealed for their sensitivity to bactericidal effect of SP ZP2. Conclusion. The synthetic ZP2 peptide is able to inhibit growth of Gram-negative bacteria, as well as exerts a bactericidal effect, thus considering it as a promising candidate for development of new effective drugs with combined immunobiological properties for combatting infectious and inflammatory conditions caused by the indicated microorganisms which show common resistance to a wide range of antimicrobial drugs used in clinical practice.

Russian Journal of Immunology. 2021;24(2):221-228
pages 221-228 views

In vitro induction of regenerative and osteogenic activity of PDLSC cells

Sukhovey Y.G., Kostolomova E.G., Unger I.G., Akuneeva T.V.


The restoration of damaged tissues in periodontal diseases is not only a medical problem, but also a social one. Periodontal diseases often entail the loss of a large number of teeth (more than with any other disease of the dentition), as a result, a violation of the act of chewing and speech, a negative effect on the general condition of the body and, as a result, a decrease in the quality of human life. Purpose of the study: to study the regenerative activity of a lyophilized extract of a chicken embryo of various concentrations in the composition of hyaluronic acid in relation to the culture of PDLSC cells in an in vitro experiment. Comparison groups: A solution of 1% unmodified hyaluronic acid containing lyophilized chicken embryo extract in three concentrations: 300 μg/ml, 150 μg/ml, 75 μg/ml. As a control, 1% solution of unmodified hyaluronic acid. Hyaluronic acid is a natural substance that is an important component of the extracellular matrix as a mineralized and non-mineralized tissue. Its use attracts the attention of specialists as an object capable of acquiring new properties with its various modifications. In our laboratory studies, stem cells from a culture of human periodontal disease were used. Periodontitis stem cells (PDLSC periodontal ligament) were discovered in 2004. Cell adhesion and tissue penetration were investigated by impedimetry. Analysis to assess cell viability was carried out using a solution containing a water-soluble tetrazolium salt. Differentiation of osteogenic direction without induction was measured three weeks after dilution of stem cells in traditional culture medium. Staining was carried out according to the Koss method. To assess mineralization, cells were stained with alizarin red, followed by assessment of calcium deposition in them. It was found that the resulting PDLSC cell population during the experiment was heterogeneous and showed healthy fibroblast morphology in all three study groups. Lyophilized extract of chicken embryo as part of a preparation based on hyaluronic acid does not significantly affect the survival and proliferation of PDLSC cells, however, at high concentrations (150 μg/ml and 300 μg/ml) it induces osteogenic activity of cells, increases mineralization without causing calcium deposition, which indicates regenerative activity. Osteogenic transdifferentiation is an attractive way to create cells of osteogenic cellular origin. The results of our study show that they can be used to model bone diseases.

Russian Journal of Immunology. 2021;24(2):229-236
pages 229-236 views

Role of S100b protein and BDNF in immunopathogenesis of neuroinflammation in the patients with induced immunocompromised state

Knysh S.V., Markelova E.V., Zenina A.A., Kostushko A.V., Fedyanina L.N.


Post-operation cognitive dysfunctions, related to mild manifestations of neurological pathology occuring after surgery, represent an important challenge for both fundamental and clinical medicine. The underlying phenomena of neuroinflammation are still poorly understood and discutable. The studies in immunopathogenesis of neuroinflammation may help in understanding the development of pathology and suggest ways to solve this problem. A special role for implementation of this process belongs to neuropeptides. Objective: to characterize immunopathogenesis of neuroinflammation in the individuals with immunocompromised state induced during aorto-coronary bypass surgery. A total of 110 patients with ischemic heart disease who underwent myocardial revascularization under extracorporeal circulation conditions were examined. Pathological neuroinflammation was recognized by development of postoperative cognitive dysfunction based on changing results of the cognitive examination (Montreal scale, MoCa test). The patients were tested before intervention, and on day 7 after surgery. According to the test results, all the patients were divided into 2 groups: (1) without signs of postoperative cognitive dysfunction and (2) with the mentioned signs. Venous blood was collected four times, on the day of surgery before the intervention, immediately after the surgery, 24 hours later, and on the 7th day after surgery. S100b and BDNF contents were determined in the venous blood serum. The baseline level of the S100b protein in the study groups did not differ from the reference values. After surgery, an increase in S100b was observed in both groups, but in patients with emerging postoperative cognitive dysfunction, these indexes were significantly higher. Despite a tendency for stabilization, the increased level of neuropeptides persisted 24 hours after the surgery; on the 7th day, the index was within the reference values, but in the 1st group it was lower than in the 2nd group. Prior to the operation, the BDNF level was significantly higher in the patients from group 1, compared with group 2. In the second group, the index was lower than the values obtained from the control group volunteers. After surgery, the values of BDNF in blood serum showed some differences: immediately after surgery, the level of neuropeptide was higher in patients without cognitive dysfunction. On day 7, the level of BDNF was within reference values, but in patients from group 2, the values were higher than in group 1.

Russian Journal of Immunology. 2021;24(2):237-242
pages 237-242 views

Transformation of the T-lymphocyte profile in peripheral blood upon resection of tuberculous lung granuloma

Berdyugina O.V.


It is known that subpopulations of peripheral blood T-lymphocytes in patients with tuberculous lung granuloma may be used for determining patient's condition and monitoring pathological process. The aim of this study was to monitor the number and functional state of peripheral blood T-lymphocytes upon resection of tuberculous granuloma of the lungs. Twenty-two patients with tuberculous lung granuloma were examined, including 12 men (54.5%) and 10 women (45.5%). The patients with immune-dependent diseases and HIV were not eligible for the study. All these patients underwent resection of the granuloma by minimally invasive method. The blood samples were examined twice: before and 5-7 days after surgery, i.e., total T-cell numbers, (CD45+CD3+), their subpopulations were also determined (CD3+CD4+, CD3+CD8+, CD3+CD4-CD8-, CD3+CD4+CD8+, CD3brightCD4-, CD3+CD16+CD56+, CD3+HLA-DR+, CD3+CD25+, CD3+CD25+HLA-DR+), as well as CD3+CD25+/CD3+HLA-DR+ ratio, performed by means of Coulter Epicx XL instrument (Beckman Coulter, USA). In addition, the changes in B- and NK-cell numbers were studied. Due to small size of the groups, distribution was considered abnormal, and the Wilcoxon Matched-Pair Test was used. The differences were considered significant at pw < 0.05. The statistical programs Statistica v. (StatSoft, USA) and GraphPad Prism v. 8.0.2 (GraphPad Software, USA) were used in the work. Resection of tuberculous granuloma of the lungs was followed by redistribution of T-, B-, and NK-cell numbers, as compared with preoperative level. The B-cell population increased by 35.8%, whereas the number of NK-cells decreased by 18.9% (pw < 0.05). Despite such significant changes in two other populations, total number of T-lymphocytes decreased only slightly: the changes ranged within 3.2% for absolute counts, and 3.8% (pw < 0.05) for relative values. The changes of T-cell subpopulation profile were different. During the postsurgical period, a 4.8% decrease in the CD3+CD4+-cell number was observed, whereas reduction in CD3+CD8+ subpopulation was more significant, and amounted to 26.2% (pw < 0.05). The number of doublepositive cells increased by 13.6% (pw < 0.05); γσT-cell counts decreased by 20.8% (pw < 0.05), and absolute number of TNK-cells increased by 32.4%. A decreased number of γσT-cells in early postoperative period could be considered a favorable criterion for assessing the patient's condition, since some previous studies provide information about increased number of these cells in patients with tuberculous granuloma. Increased HLA-DR expression on T-lymphocytes may be another favorable factor, since the patients with tuberculous granuloma are known to have lesser amounts of such cells in peripheral blood than healthy persons. In general, a search for expression of activation molecules showed that there is a decrease of T-cells expressing CD25+, and an increased number of CD3+HLA-DR+-cells. The T-lymphocyte population co-expressing both CD25+ and HLA-DR+ shifts to the normal ranges after the surgery. The obtained data are supposed for monitoring results of treatment in the patients.

Russian Journal of Immunology. 2021;24(2):243-248
pages 243-248 views

Cytokine-neuroendocrine peripheral signature in the context of the “accelerated ageing” phenomenon in autism spectrum and schizophrenia spectrum disorders

Filippova Y.Y., Burmistrova A.L.


Currently, differentiation between autism spectrum disorders and schizophrenia spectrum disorders in children is a difficult task, because it relies mainly on behavioral and symptomatic characteristics, since these disorders are highly similar. We have previously demonstrated that peripheral indexes of immune and neuroendocrine systems, which we combined into cytokine-neuroendocrine signature, may reflect distinct clinical phenotypes of autism and schizophrenia spectrum disorders. Moreover, a number of researchers discovered the “accelerated ageing” phenomenon in the persons with schizophrenia, which includes deficiencies of cognitive functions and performance as the main symptoms. Here we carried out a search for biological markers of the “accelerated ageing” phenomenon in children with autistic conditions and schizophrenia spectrum disorders. Our aim was to assess the opportunity of applying the cytokine-neuroendocrine signature as biological evidence of “accelerated ageing” phenomenon in children with autism and schizophrenia spectrum disorders, which could be potentially useful for differential diagnosis of these disorders.

Thirteen parameters of the cytokine-neuroendocrine signature were assessed in blood plasma using ELISA method in 82 children with autism, 9 children with schizophrenia, 45 normally developing children, 25 subjects in their reproductive age, and 39 elderly persons: cytokines (IL-6, IL- 1β, IFNγ, TNFα, IL-10, IL-4) and neurohormones (oxytocin, dopamine, adrenaline, noradrenaline, adrenocorticotropic hormone, cortisol, and serotonin). The nonlinear principal component analysis (CATPCA algorithm) was used to assess the variants of cytokine-neuroendocrine signature for different diagnostic categories, i.e., “autism spectrum disorders”, “schizophrenia spectrum disorders”, and “healthy ageing”.

The “healthy ageing” variant of cytokine-neuroendocrine signature presented a classic phenomenon, referred to as immune senescence presented by pro-inflammatory age-related cytokines — IL-6, IL- 1β, IFNγ. Only the “schizophrenia spectrum disorders” variant of the cytokine-neuroendocrine signature, unlike all the other signature variants, demonstrated high-level similarity with the “healthy ageing” variant (differing in 2 out of 13 indexes): lower levels of IL- 1β and IFNγ, at the same level of IL-6 “gerontological cytokine” index.

Evaluation of the cytokine-neuroendocrine signature can be used for differentiation between autistic disorders and schizophrenia spectrum disorders, including predictive diagnostics in children with autism, thus enabling group selection of children at risk for later conversion to schizophrenia.

Russian Journal of Immunology. 2021;24(2):249-256
pages 249-256 views

Anticytokine activity and the ability to produce cytokine-like substances of staphylococci isolated from the prostatic secretions in men with symptoms of urogenital infection

Kartashova O.L., Pashkova T.M., Pashinina O.A., Morozova N.V.


To be causal agents for infectious and inflammatory diseases, the microorganisms should have a pathogenic potential, the framework of which is based on persistence factors that ensure their protection from various effectors of the host's immunity. There is evidence that some microorganisms can have anti-cytokine activity (ACA), and at the same time, many microorganisms many synthesize cytokine-like substances, thus, in fact, exhibit cytokine activity. The aim of the present study was to establish the presence of cytokine-like substances and anti-cytokine activity in Staphylococcus strains isolated from prostate secretions in men with symptoms of urogenital infection. This study involved 24 clinical isolates of different Staphylococci types isolated from the prostate gland secretions in men with symptoms of urogenital infection. Ability of staphylococci to cause changes in the IL-4, IL-6, IL-8, IL-10 and TNFα contents was carried out by co-incubating bacteria suspended in normal saline solution together with cytokine solutions, at a ratio of 1:1 (experimental samples). As a control, a solution of cytokines supplemented by normal saline was used. Concentration of cytokines in experimental and control samples was determined by enzyme immunoassay. Anti-cytokine activity (ACA) was expressed as a percentage (%) of cytokine inactivation in experimental mixtures compared to the controls, cytokine activity, as % of cytokine production in the experimental samples compared to the control.

The studied staphylococci showed anti-cytokine activity against IL-4 and IL-10, and exhibited ability to produce IL-6 and TNF. At the same time, among staphylococci, there were strains both showing anti-cytokine activity against IL-8, and producing this cytokine.

Among different staphylococcal species isolated from prostate secretions in men with symptoms of urogenital infection, the presence of anti-cytokine activity and the ability to produce cytokines was detected; their production (presence and level) was characterized by intraspecific variability. S. xylosus proved to be the most active cytokine producer, which also have the highest anti-cytokine activity against IL-4.

Russian Journal of Immunology. 2021;24(2):257-260
pages 257-260 views

Dynamics of cellular factors of innate immunity and antioxidant protection in the patients with chronic recurrent vulvovaginal candidosis upon exposure to cavitated physiological solution

Sorokin Y.A., Gizinger O.A.


Dysfunction of colonization resistance factors at the mucous membranes of genitourinary system during adhesion of Candida fungi is a common pathogenic situation leading to impairment of the mucous membrane regeneration, unsuccessful attempts at in vitro fertilization (IVF), increased risk of complications during gestation. The aim of present study was to substantiate the opportunity of complex therapy for chronic vulvovaginal candidiasis using cavitated saline and usage suppositories with alpha-2b human recombinant interferon a (Viferon LLC, Russia). The study involved 90 women at the age of25.45±7.16 years, with clinically and laboratory confirmed recurrencies of chronic vulvovaginal candidiasis observed, on average, 5.55±0.45 times. Group 1 consisted of 30 women with C. albicans infection of urogenital tract, at the mean age of 27.11±1.52 years, who received therapy with fluconazole (150 mg once every 7 days). Group 2 consisted of 30 women with C. albicans infection of mucous membranes, mean age 27.31±1.99 years, treated with Fluconazole (150 mg 1 time in 7 days, and 5-min. irrigation of vagina with ultrasonicated saline solution, 7 days; Low-intensity ultrasound exposure was performed using the Fotek AK100-25 hardware complex, ultrasonic vibration frequency of 25 kHz (Yekaterinburg, Russia). Group 3 consisted of 30 women whose treatment included Fluconazole (150 mg once every 6 days), irrigation of mucous membranes with ultrasonicated cavitated solution and subsequent administration of suppositories containing Viferon® 500,000 IU (1 suppository daily for 7 days). Biochemical studies included determination of isopropanol- and heptane-soluble primary, secondary end products of lipid peroxidation. Activity of the antioxidant system was studied by determination of superoxide dismutase, glutathione peroxidase, ceruloplasmin, vitamin C. Qualitative and quantitative characteristics of neutrophilic granulocytes from vaginal secretions were evaluated by testing phagocytic activity, reduction of nitroblue tetrazolium compound to diformazan. The studies of IL-8, TNFα, IL-2, and IL-10 cytokines were carried out in cell-free supernates of vaginal secretions by ELISA test systems (LLC “Vector Best”, Novosibirsk). Results: upon therapeutic exposure to low-frequency ultrasonic cavitation combined with antioxidant drug, we have observed a decreased local concentration of pro-inflammatory cytokines, the balance of lipid peroxidationantioxidant defense system was restored, and functional metabolic status of the phagocytes in vaginal secretions was normalized.

Russian Journal of Immunology. 2021;24(2):261-268
pages 261-268 views

Visualization of B10 regulatory cells in peripheral blood in physiological pregnancy

Ziganshina M.M., Khaidukov S.V.


The current trend in studies of the B-cell immunity is the study of small subpopulations of cells. It was found that a minor subpopulation of IL-10 producing B-cells (B10-reg cells) has the properties of limiting excessive reactions of the innate and adaptive immune response. Their regulatory and pathogenetic effect has been shown in various physiological and pathophysiological conditions, in particular in the pathological pregnancies.

Due to the low content of B10-reg cells in the blood (up to 1%) and the difficulties of visualizing flow cytometry data, a previously developed method based on prolonged stimulation of peripheral blood cells with a combination of factors ((CD40L+CpG) and PMA) causing cell activation, proliferation and maturation, allows visualization of the enriched fraction of B10-reg cells (B10 cells + pro-B10 cells), the content of which exceeds 5%. The aim of this study was to obtain a stimulated ex vivo population of B10 cells + pro-B10 cells from the peripheral blood of patients with physiological pregnancy and to develop an optimal strategy for gating B10-reg cells for their visualization.

Materials and methods: in patients with physiological pregnancy in the third trimester, peripheral blood was taken. The cells were stimulated according to two protocols. First protocol: short (5 hours) stimulation of whole blood cells under sterile conditions with mixture of PMA + ionomycin + brefeldin A. The second protocol: long-term (48 hours) stimulation of the isolated mononuclear fraction under sterile conditions with a mixture (CD40L + CpG) with the addition of PMA + ionomycin + brefeldin A during the last 5 hours. Cells were stained for surface markers (CD45, CD19, CD24, CD27, CD38) and the intracellular content of IL-10. Sample analysis was performed on a Navios™ flow cytometer.

Results: a five-color cytometric analysis was performed and a sequential gating strategy was developed based on the isolation of the gate by lymphocytes (marker CD45); restriction of B-lymphocytes (marker CD19); isolating a subpopulation of B cells expressing the CD24 marker; limiting the two required subpopulations of B10-reg cells for CD27 and CD38: CD19+CD24hiCD27+IL-10+ and CD19+CD24hiCD38hiIL-10+. Stimulation of cells based on the first protocol allows visualizing up to 1% of both subpopulations, and based on the second protocol - about 10%. The method opens up prospects for fundamental research of B10-reg cells during pregnancy. The detectable amounts of an enriched population of B10-reg cells can be of diagnostic and prognostic value in the clinic for idiopathic obstetric complications.

Russian Journal of Immunology. 2021;24(2):269-274
pages 269-274 views

Cytokine profile in the subjects after long-term in utero and postnatal exposure to chronic irradiation

Kodintseva E.A., Akleyev A.A., Blinova Е.A., Akleyev A.V.


Persons exposed to ionizing radiation in utero and in early childhood constitute a risk group for the development of long-term stochastic consequences of irradiation. The imbalance of cytokines at the long terms after irradiation could be considered a carcinogenic triggering factor in subjects previously irradiated in utero and in early childhood, thus determining relevance of the study. The aim of the present study was to assess the levels of serum cytokines in native residents of coastal villages at the Techa River, whose chronic irradiation had been begun antenatally and to study probable interrelatirons between the detected changes, radiation and non-radiation factors at long terms after the exposure was begun. The main group included 61 persons from the Techa River Cohort who were born in 1950-1960, whose irradiation was begun in utero, being continued over the early postnatal period. For patients from the main group, the mean dose of antenatal radiation was calculated 74.7 mGy for red bone marrow as of, the mean dose of postnatal irradiation was calculated for red bone marrow as 537.5 mGy, and the median age of patients was 64.0 years. The comparison group (90 nonirradiated persons) was comparable to the main group in terms of age, gender, ethnicity and socio-economic status. The median levels for IL-2 in the main group were 1.37 pg/ml; in the comparison group, 2.70 pg/ml, p = 0.020; for IL-10, 4.53 pg/ml versus 7.58 pg/ml, p = 0.030 respectively; for GM-CSF, 0.39 pg/ml in the subjects who were irradiated in utero and in the early postnatal period versus 0.86 pg/ml in non-irradiated persons, p = 0.040. The median serum concentrations of IL-1β, IL-1α, IL-1(ra), IL-4, IL-6, IL-8, G-CSF, TNFα, IFNα, IFNγ in the study group did not show differences from the values in a group of non-irradiated persons. The decrease of the serum IL-2, IL-10 and GM-CSF levels in the persons of the main group did not depend on the dose of antenatal irradiation to red bone marrow, and on the radiation dose to red bone marrow received during the postnatal period of ontogenesis. In the main group, there was a moderate inverse relationship between the serum IL-10 level and age at the time of examination (SR = -0.53, p < 0.001). Serum concentrations of IL-2 and IL-10 in the people from comparison group showed a moderate positive correlation with their present age (SR = 0.47, p < 0.001 and SR = 0.42, p < 0.001 respectively).

Russian Journal of Immunology. 2021;24(2):275-282
pages 275-282 views

Age-dependent patterns of the baseline cytokine levels in unstimulated peripheral blood samples

Lagereva Y.G., Palitsyna O.V., Ischenko N.V.


The cytokine levels in unstimulated peripheral blood samples differ in relation to age, living conditions (environmental situation, state of medical care, quality of life, etc). Thus, the essential task is to determine normal levels of peripheral blood cytokines for different age groups with respect to regional characteristics. This study aimed to investigate age-associated patterns of the baseline levels for some cytokines in unstimulated peripheral blood samples in healthy children and adults. The studies were carried out at the Laboratory of Clinical Immunology (Clinical and Diagnostic Centre, Yekaterinburg). Peripheral blood samples were taken in 149 healthy children aged from 7 months to 18 years. Group 1 was 7 to 12 months old (29 infants); group 2 was 1 to 3 years old (33 children); group 3 was 4 to 7 years old (29 children); 4th group was 8 to 14 years old (26 children); 5th group, 15 to 18 years old (32 children); and 6th group included 42 adults (19 to 45 years old). The concentrations of TNFα, IFNγ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-17, and IL-18 in blood serum were determined by common ELISA technique using Vector-Best kits (Novosibirsk, Russia). Analysis of the data made it possible to identify various patterns of age-dependent changes of baseline levels for the determined cytokines: IL-2 showed an increase in concentration by the age of 1-3 years, stable contents up to 18 years, and decreased concentrations in adulthood. IFNγ was at zero levels in children and adolescents, followed by increased levels in adults. IL-4 and IL-6 showed consistently low levels in children and adolescents, with a decrease to zero levels in adult subjects. IL-18, TNFα were at maximal levels at the age of below 7 years followed by a decrease in concentration; IL-1β, IL-17 exhibited bimodal changes, with maximal contents at the age of 1-3 years, and repeated increase in adult age. Nearly stable baseline concentration was noted for IL-10, with a slight increase at 1-3 years, like as for IL-8, with a moderate increase at 15-18 years. For majority of analyzed cytokines, the maximal baseline contents in unstimulated peripheral blood samples were found in children aged 1-3 years.

Russian Journal of Immunology. 2021;24(2):283-290
pages 283-290 views

Relationships between spermogram indexes in males with chronic bacterial prostatitis and the levels of local antimicrobial factors

Bekpergenova A.V., Bondarenko T.A., Perunova N.B., Chelpachenko O.E., Nikiforov I.A.


Sperm quality can be directly or indirectly affected in chronic bacterial prostatitis, due to protein components of mucosal immunity, including lactoferrin, secretory immunoglobulin A, and lysozyme. Therefore, a search for relationships between spermogram indices and levels of antimicrobial factors in sperm plasma presents an urgent task in healthy men and patients with chronic bacterial prostatitis. The present paper contains the results of a study of sperm samples from 72 men aged 20 to 45 years. Samples were collected in sterile containers by masturbation after a minimum abstinence period of 3-5 days. None of the patients had previously taken antibiotics. Patients with sexually transmitted infections were excluded from the study. The patients were divided into two groups — conditionally healthy males (n = 30) and patients with chronic bacterial prostatitis (n = 42). Seminal plasma was obtained by two-stage centrifugation of ejaculate samples (1000-3000 rpm) for 30 minutes. Evaluation of lactoferrin and secretory immunoglobulin A levels in seminal plasma was carried out by ELISA technique (Vector-Best, Novosibirsk), lysozyme — by turbidimetric method. The results were recorded with Multiscan Labsystems photometer (Finland) at 492-nm wavelength. Statistical evaluation of the obtained data was carried out using Statistica 10 package (StatSoft, USA). To assess the kind of relationships between the studied parameters, the Spearman's rank correlation quotient was applied. When using the Spearman rankcorrelation approach, the strength of the relationships between the features was assessed, considering the values of < 0.3 as a weak connection; quotient levels of 0.4 to 0.7 as indexes of moderate relationships, and values of 0.7 and more, as high connection indexes.

We have found that an increased level of lactoferrin in some cases of leukocytospermia, oligospermia, and asthenospermia is accompanied by decreased number of leukocytes, increased sperm motility, preservation of sperm morphology, thus suggesting improvement of the sperm quality. Considering universal biological properties of lactoferrin and its receptors, an increased number of positive correlations between level lactoferrin level and spermogram parameters was shown in the patients with chronic bacterial prostatitis, as compared with healthy males, thus presuming high diagnostic value of this marker. Hence, diagnostic value of spermogram parameters is determined not only by their quantitative values, but also by the types of correlations with antimicrobial factor levels, in particular, with those of lactoferrin, secretory immunoglobulin A and lysozyme.

Russian Journal of Immunology. 2021;24(2):291-296
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Local concentrations of secretory immunoglobulin a in patients with adenoiditis, rhinosinusitis and exacerbation of chronic purulent otitis media when using physiatric methods in complex therapy

Korkmazov M.Y., Dubinets I.D., Lengina M.A., Solodovnic A.V.


Improvement of therapeutic tools is one of topical issues in modern otorhinolaryngology. The article is devoted to the study of the stimulating effect of physical therapy on the concentrations of secretory IgA in nasal secretions, from the entries of auditory tubes and nasopharynx in the patients suffering from adenoiditis, rhinosinusitis, and exacerbation of chronic otitis media. The increase in clinical and immunological efficiency when using low-frequency ultrasonic cavitation therapy in combination with photochromotherapy in combined treatment of these conditions was evaluated in dynamics by measuring concentrations of secretory immunoglobulin levels in nasal secretions using enzyme immunoassay technique (Vector-Best, Russia). The purpose of this study was to improve clinical and immunological effectiveness of treatment of patients with chronic adenoiditis, sinusitis and exacerbation of chronic purulent otitis media when using low-frequency ultrasound cavitation and photochromotherapy in complex therapy. The study involved 54 patients. The control group consisted of 25 patients who received conventional treatment (topical and systemic antibacterial and anti-inflammatory drugs, unloading and elimination procedures, symptomatic and restorative therapy) and the main group of 29 people who, in addition to etiotropic therapy, received low-frequency ultrasound cavitation irrigation of the nasal cavity followed by a course of photochromotherapy. Evaluation of the effectiveness of treatment was carried out by studying the barrier state of the nasal mucosa by the levels of secretory IgA concentration by the method of enzyme immunoassay. The low-frequency ultrasound cavitation in combination with photochromotherapy as an adjunct to etiotropic treatment in patients with adenoiditis, rhinosinusitis and exacerbation of chronic otitis media promotes an earlier increase in secretory IgA concentration of nasal mucosa and entry of the auditory tube when compared to standard treatment methods. The trend to IgA increase by 14 days reached maximal values and was slightly decreased month later. An early increase in the concentration of secretory IgA in nasal secretions and the entry of auditory tube, and positive clinical results of treatment are observed when low-frequency ultrasound cavitation is used in etiotropic therapy in combination with photochromotherapy in patients with adenoiditis, rhinosinusitis and exacerbation of chronic otitis media. The results substantiate the opportunity of introducing these physical methods of physical therapy into complex etiotropic therapy, as a non-invasive and effective method.

Russian Journal of Immunology. 2021;24(2):297-304
pages 297-304 views

Studies of immunological and microbiological efficiency of the therapy of curcumin and methionine in the developed capsules

Khisamova A.A., Gizinger O.A., Kornova N.V., Zyryanova K.S., Korkmazov A.M., Beloshangin A.S.


Curcumin is a polyphenolic compound, the main component of the Curcuma longa rhizome. Recently, there is growing interest to studies of this new, inexpensive and safe substance that may be used to treat various diseases. Curcumin is widely used in medicine due to its therapeutic efficacy and safety. Its usage in therapeutic practice as a dietary supplement has shown that curcumin exhibits antioxidant and immunomodulatory effects. Data from clinical studies have shown its pharmacokinetic, pharmacodynamic profile and potential use of curcumin in humans for treatment of various diseases, even at the early stages of treatment. Purpose of the study: analysis of immunomodulatory and microbiological properties when using the original grained preparation containing curcumin and methionine. It is suggested to have a regulatory effect by modulating microbial richness, diversity and composition of intestinal microflora. Analysis of immunomodulatory and microbiological activity of the components in the developed dosage form of the capsules with curcumin and methionine was carried out in vitro and in vivo. Functional and metabolic properties of neutrophils were determined with addition of its components, i.e., curcumin and methionine. The analysis of intensity and activity of neutrophils was carried out using the NBT-test. The analysis showed that simultaneous incubation of peripheral blood neutrophils from ICR (CD-1) mice with curcumin and methionine leads to an increase in spontaneous and induced NBT-reducing activity, an increase in the functional reserve and phagocytic activity of peripheral blood neutrophils in mice. A study of the modulating effects of oral intake of curcumin and methionine, which are part of the dosage form, on the intestinal microbiota of ICR (CD-1) mice was carried out. It was found that curcumin, together with methionine, affects the number of some representative families of intestinal microbial communities: in total, there were 640 common operating taxonomic units. between the curcumin-methionine and control groups, 65 were unique in the curcumin-methionine group and 93 in the control group. Given the direct link between gut microbiota and certain diseases, these results may help to interpret therapeutic benefits of curcumin with methionine. The results of the study showed that the developed dosage form, which contains curcumin and methionine, shows antioxidant and immunomodulatory effects, which can be potentially used to treat diseases associated with the effects of oxidative stress on the organism.

Russian Journal of Immunology. 2021;24(2):305-310
pages 305-310 views

Glucose uptake by CD4+Т cell subsets in HIV-infected patients receiving highly active antiretroviral therapy (haart)

Korolevskaya L.B., Saidakova E.V., Vlasova V.V., Shmagel K.V.


Metabolism of glucose, a universal biosynthetic substrate in CD4+T lymphocytes, is increased in HIV-infection. However, the issue of heterogeneity in glucose consumption by the CD4+T cell subsets remains unresolved. The aim of this work was to assess glucose uptake by resting and cycling naive and memory CD4+T cells in HIV-infected patients receiving antiretroviral therapy. We examined 47 subjects, 26 of whom were infected with HIV and were administered antiretroviral drugs. The following indices were determined: the number of CD4+T lymphocytes, size of naive (CD4+CD45R0-) and memory (CD4+CD45R0+) T cell subsets, frequencies of resting (CD71-) and cycling (CD71+) cell forms in each subset, the glucose fluorescent analog (2-NBDG) uptake by various CD4+T cell subsets. It was shown that, regardless of HIV status, the CD4+T cells are active glucose consumers. Cycling lymphocytes, compared with resting cells, uptake more biosynthetic substrate. We have revealed a trend for increased glucose uptake in HIV-infected patients when compared with healthy individuals. Memory cells, in comparison with naive lymphocytes, consume the substrate more actively, independent on HIV status. At the same time, naive CD4+T lymphocytes of HIV-infected individuals capture more glucose than the corresponding cell subset in non-infected donors. Cycling naive CD4+T lymphocytes of HIV-positive subjects are more active consumers of glucose than the analogues in healthy subjects. No differences were found between HIV-positive and HIV-negative groups for intensity of substrate consumption by the cycling memory CD4+T cells. Thus, in treated HIV-infected patients, CD4+T cells seem to uptake more glucose than similar cell subpopulations in healthy people, which, apparently, is mediated by the activity of resting naive lymphocytes. The data obtained indicate that metabolic characteristics in resting T-cells are instable and may change depending on the substrate availability.

Russian Journal of Immunology. 2021;24(2):311-316
pages 311-316 views

Comparative analysis of treatment efficiency in chronic periodontitis using experimental therapies with peptide-containing preparation and polyoxidonium

Sarkisyan N.G., Chumakov N.S., Khlystova K.A., Drozdova L.I., Yuffa E.P.


In this article, a comparative analysis of two experimental methods for treatment of chronic parodontitis was carried out, using a locally applied composition of silicon glycerohydrogel — peptide, and local injections of “Polyoxidonium” drug. The aim of this study was to compare efficiency of the two experimental methods for treatment of chronic periodontitis. Materials and methods: Using the facilities at the Institute of Immunology and Physiology (Yekaterinburg), a model of chronic inflammation of periodontal tissues was developed in Wistar rats, further treated by two experimental methods, i.e., (1) applying a composition containing organosilicon glycerohydrogel and a synthetic peptide, or (2) injections of “Polyoxidonium” drug directly into the foci of inflammation. Subsequently, a comparative evaluation of their efficiency was performed, including comparisons with control groups. The latters were treated by topical use of organosilicon glycerohydrogel (Group 1), and the local application of “Metrogyl Denta” gel (Group 2). Upon completion of the treatment, clinical and histological data were evaluated and compared. As based on the data obtained, we have found that all these drugs exerted favorable effect on the tissue regeneration, and led to reduced intensity of inflammatory processes. We have revealed also that the organosilicon glycerohydrogel — peptide composition provided a faster effect, due to the special characteristics of its components. The hydrogel, which has transcutaneous activity and plays a role of conductor substance, promotes faster penetration of the peptide into the tissues, thus allowing the peptide substance for more pronounced, complex antimicrobial and regenerative effect upon various pathogenetic components of chronic perirodontitis. If compared with the groups treated by glycerohydrogel silicon and “Polyoxidonium”, the terms of clinical improvement in the glycerohydrogel-peptide group were found to be increased by 57%, and appropriate indexes in the group treated with “Metrogyl Denta” improved by about 15%. The results of histological examination have confirmed normalization of the local tissue structure, as well as decreased inflammatory response observed for all the groups. Of particular interest, regeneration terms in the glycerohydrogel-peptide group were shorter (16 to 20 days) than in other groups (from 20 days), thus suggesting a more pronounced effect of this composition when compared to other treatments. Due to the presence of antimicrobial peptides in formulation of the glycerohydrogel-peptide composition, both pathogenetic and etiological links of the disease are affected, thus being important for development of integral approach to the treatment of chronic periodontitis.

Russian Journal of Immunology. 2021;24(2):317-322
pages 317-322 views

Effect of local immunotherapy on the indexes of mucosal immunity in patients with surgical dental pathology

Latyushina L.S., Malysheva L.Y., Piotrovich A.V., Berezhnaya E.S.


Currently, inflammatory processes and injuries of maxillofacial region, occupy leading positions in the overall structure of surgical morbidity. Restoration of dentition integrity is based on intraosseous dental implants, being a widespread method of treatment in dentistry. The aim of the present work was to perform a comprehensive analysis of cytokine contents (IL- 1β, IL-1ra, IL-2, IL-8, IL-10, TNFα, IFNγ) in maxillary sinus discharges and mixed oral fluid in the patients with surgical dental pathology (chronic odontogenic maxillary sinusitis, complicated by mandibular fracture, as well as during simultaneous dental implantation) and to assess the effect of locally applied immunotherapy on the parameters studied. As a part of the two-stage study, 218 people were comprehensively examined, divided into groups depending on the type of disorder and methods of treatment. The patients with chronic odontogenic sinusitis (J 32.0, n = 44) were treated with topical immunotherapy with Cycloferon (meglumine acridonacetate), patients with a mandibular fracture (S 02.6, n = 70) and with a dental implant (K 08.1, n = 70) received Betaleucine (recombinant IL-1β). At the first stage of the study, examination of all the patients with surgical dental pathology showed immunological predictors of low-efficient acute inflammatory response associated with imbalance of the pro- and anti-inflammatory cytokine system. These predictors were primarily found in the patients with chronic odontogenic sinusitis, i.e., a decrease in IL-1β at high IL-1ra values in the maxillary sinus secretions. In the patients with complicated mandibular fractures, there was a pronounced dynamic decrease in IL-1β with consistently low values of TNFα in mixed saliva. There were distinct dynamic features of IL-1β, IL-8, and IL-1ra levels in oral fluid of the subjects undergoing implantological procedures. The second stage of the study, associated with the local use of Cycloferon and Betaleucine, and analysis of the data revealed different effects of the drugs on the studied parameters, presuming an integral effect of topical immunotherapy, i.e., some leveling of the previously detected imbalance of the cytokine system, due to normalization of their concentrations. Hence, the results obtained may indicate that the patients with surgical dental diseases treated with local immunotherapy had a higher adaptive potential of the local mucosal immunity.

Russian Journal of Immunology. 2021;24(2):323-330
pages 323-330 views

Immunotropic effect of medicinal plants with different microelemental composition

Ivanova E.V., Voronkova I.P., Bondarenko A.I., Tarenkova I.V.


Increasing use of raw materials from medicinal plant is due to the fact that the natural compounds may be preferred to synthetic ones by, generally, low toxicity, absence of side effects and addiction. Over recent decades, much attention has been paid to the study of immunotropic and antioxidant effect of medicinal plants, as well as to analysis of their trace element composition, since the action of the main biologically active substances is often manifested in combination with the natural mineral composition of the plant. The aim of the present work was to assess immunotropic effect of aqueous extracts of officinal medicinal raw materials and to analyze their trace element composition. The objects of the study were water extracts (1:10) from industrial samples of 11 types of raw materials (black currant leaves (Ribes nigrum L.), yarrow herb (Achillea millefolium L.), licorice roots (Glycyrrhiza uralensis Fisch.), Flowers of the sand immortelle (Helichrysum arenarium (L.) Moench), St. John's wort (Nurericum perforatum), wild strawberry leaves (Fragaria vesca L.), wild bird cherry (Padus avium Mill.), blood-red hawthorn (Crataegus sangunea Mill.) (Tanacetum flowers) vulgare L.), common chicory root (Cich rium ntybus Fisch.) and oat grass (Avena sativa L.) supplied to the network of pharmacies in Orenburg. Immunotropic effects of raw plant materials were evaluated in a model of adaptive immunity cells, by their ability to induce production of cytokines: IL-1β, TNFα and IL-10. The level of cytokines (IL-1β, TNFα, and IL-10) were assessed using ELISA (Cytokin, St. Petersburg, Russia). Elemental composition of raw material samples was determined by mass spectrometry with inductively coupled plasma using the ELAN-DRC-e device (PerkinElmer SCIEX, USA). Analysis of immunotropic activity of aqueous extracts from officinal medicinal plants showed that the majority of plant raw materials are characterized by a suppressive effect both upon pro-inflammatory mediators (TNFα, IL-1β), and the main anti-inflammatory cytokine (IL-10). In contrast, aqueous extracts of yarrow, oats and chicory were characterized by a selective immunomodulatory effect aimed at suppressing only inflammatory mediators. A tendency has been established for a significant accumulation of such biologically important trace elements as Zn, Mn, Fe and Cu in medicinal plants, which can determine their biological activity, and allows them to be considered as promising components at the stage of developing drugs that both exert immunotropic effect, and are a source of microelements.

Russian Journal of Immunology. 2021;24(2):331-336
pages 331-336 views

Dynamics of antioxidant protection factors, cytokines in patients with chronic streptodermia, correction methods

Gizinger O.A., Laknitskaya A.O., Ziganshin O.R.


Certain relationships exist between different biological systems, i.e., lipid peroxidation/ antioxidant defense system; changing balance of Th1/Th2 immune response and cytokines; a decrease of interleukin 2 (IL-2) concentration; lower number of IL-2 receptors on CD25+ lymphocytes, impaired activity of protective antioxidant enzymes (superoxide dismutase and catalase); decreased general antioxidant status. These connections justify an approach to treatment of chronic streptoderma, using combined effects upon all components of the pathological process, by restoring the cytokine balance, eliminating oxidative stress and chronic inflammatory foci on the skin surface. The study group consisted of 50 male patients with chronic streptoderma, lasting for more than 2 years. Average age of the patients was 32.25±4.23 years; for comparison, the indexes of healthy volunteers were used on the basis of informed consent. The aim of the present study was to propose a method for treating chronic streptoderma using recombinant interleukin-2 and low-intensity laser irradiation justified by assessment of immune profile, pro-oxidant and antioxidant status. Standard therapy included local treatment of the affected surface with 1% aqueous solution of chlorhexidine bigluconate; 2% ointment with mupirocin as an active substance, applied to the area of rashes for 7 days; treatment of the skin surface with a low-intensity laser at the wavelength 40 ±10 nm (blue light) using a Mustang device (Moscow, registration certificate No. RZN 2014/1410 of 06.02.2014), according to technical conditions TU 9444-005720850602008. The irradiation exposure mode corresponded to recommendations of the device manufacturer. The radiation power was 80.0±5.0 mW, applied for 10 days, along with subcutaneous administration of recombinant IL-2 (Roncoleukin®) 500,000 IU, diluted with saline at the interval of 24 hours for 5 days. Positive clinical dynamics was noted during complex treatment which involved recombinant IL-2 and low-intensity laser exposure in the patients with chronic streptoderma, according to evaluation by a visual analogue scale, normalization of the cytokine balance (decreased contents of pro-inflammatory cytokines), and normalization of antioxidant status. Thus, the use of blue light exposure at the visible range (wavelength of 450.0±10.0 nm) and subcutaneous injections of recombinant IL-2 (Roncoleukin®, 500,000 IU) is a justified method in complex therapy of chronic streptoderma.

Russian Journal of Immunology. 2021;24(2):337-342
pages 337-342 views


Памяти Анатолия Николаевича Чередеева. К 80-летию со дня рождения



Russian Journal of Immunology. 2021;24(2):343-344
pages 343-344 views

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