Vol 26, No 4 (2023)

Complex immunobiology of lymphotoxin (LTα) is due to multiple modalities of signal transduction, involving a soluble homotrimer and membrane-bound heterotrimers that engage at least three different receptors. While LTα is crucial for the formation and maintenance of secondary lymphoid organs, its overproduction is observed in autoimmune diseases such as rheumatoid arthritis and multiple sclerosis. Initially, LTα was considered pathogenic in the development of experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis, as demonstrated by the resistance of mice with genetic LTα inactivation to EAE induction. However, conflicting observations arose when EAE was induced in RAG1-deficient mice that underwent adoptive bone marrow transfer from LTα-deficient mice, thereby calling into question previous conclusions about the role of LTα in EAE development.

This study aimed to investigate the role of LTα in MOG_35-55-induced EAE using mice deficient in LTα or its membrane receptor, LTβR. LTα knockout mice used here were designed to avoid the artifact involving TNF gene downregulation in myeloid cells, which occurred in the conventional LTα knockout mice.

Surprisingly, LTα-deficient mice with normal TNF expression developed EAE clinically comparable to wild-type mice. Conversely, genetic inactivation of LTβR delayed EAE onset. However, during the later stages of the disease, LTβR deletion exacerbated clinical symptoms of EAE.

These findings demonstrate that the involvement of LTα in EAE development is more complex than previously estimated, and that LTβR exhibits diverse functions depending on the disease stage: pathogenic at the early stage and protective at the later stages of EAE.

Myeloid-derived suppressor cells (MDSCs) are of interest as key regulators of the immune response for the development and improvement of cellular technologies in biomedicine. Enhancing the suppressive activity of these cells is important for developing therapies for autoimmune diseases and miscarriages, and their suppression may be useful in the treatment of cancer, since MDSCs are known to suppress antitumor immunity. However, there is a problem that prevents the active study of MDSCs, i.e., the difficulty in obtaining sufficient numbers of this cell population. Isolation of MDSCs in cancer patients poses an ethical challenge. Moreover, these MDSC may differ in subpopulation composition and suppressive activity due to individual factors. Researchers who generate human MDSC from bone marrow cells may also face similar problems. Therefore, finding a reliable and affordable source of these cells to facilitate the study of their functions is extremely important. Attempts to obtain human MDSCs in vitro have been ongoing for a long time. GM- CSF, IL-6, IL- 1β, IL-4, PGE2, LPS, M-CSF, IFNγ are described as factors that induce the ex vivo MDSC differentiation. However, despite multiple factors used, not all protocols are clearly reproducible, leading to generation of a sufficient number of cells in the target population. Previously, we had also developed a scheme for MDSC differentiation from CD11b⁺ cells derived from human peripheral blood, which made it possible to obtain a tangible but still insufficient percentage of cells to study functional activity.

To increase the number of MDSCs in cultures, we developed a protocol aimed for differentiation of these cells from peripheral blood monocytes (CD14⁺ cells) previously transformed into PCMO (programmed cells of monocytic origin). The monocytes isolated by immunomagnetic separation were cultured in a de-differentiating medium (complete culture medium supplemented with M-CSF, IL-3 and β-mercaptoethanol) for one week. Later on, the medium was replaced by the addition of GM-CSF, being cultured for three days, followed by addition of LPS and IL-1β in order to induce suppressive activity. We have found that culturing CD14⁺ cells on a two-week schedule with prior creation of dedifferentiation conditions resulted in a slightly decreased percentage of viable cells in culture. However, there was a trend towards an increased ratio of MDSCs in culture (from an average of 34 to 40%) and an increase in their suppressive activity (arginase and IDO expression). The percentage of Arg⁺ cells increased by average of 10%, and IDO⁺ cells, by 16%. Moreover, the percentage of mature M-MDSCs was significantly (several-fold) higher when compared with differentiation protocol using CD11b⁺ cells. Hence, this method of MDSCs production enables us to increase the number of cells belonging to the conditionally “mature” monocyte subpopulation of MDSCs, as well as the percentage of functional suppressor cells in the population. The described scheme may be used to improve the quality of studies aimed at modulating MDSC functions in order to develop new therapeutic approaches.

The article deals with usage of a novel immunotropic drug composition in experimental model of periodontitis. The composition contains silicoorganic glycerogel, exhibits broad spectrum of action upon the etiological pathogen. A good therapeutic effect was revealed by clinical, laboratory and histological criteria. Histology showed absence of osteoclasts, reduced lacunas, higher density and normalization of tissue structures, recovery of microcirculation, angiogenesis and formation of granulation tissues. However, the phosphorus-to-calcium ratio in the group supplied with peroral vitamin D treatment, was characterized by significant decrease in alkaline phosphatase and inorganic phosphorus thus confirming efficiency of complex topical effect of the given composition and vitamin D.

Alteration of microbiota composition is a trigger, and, sometimes, an etiological factor in the development of chronic skin diseases, e.g., psoriasis or eczema. Recognition of microflora by keratinocytes and immune cells leads to the production of antimicrobial peptides, chemokines and growth factors, which contribute to the differentiation of T lymphocytes to autoaggressive effector cells of Th1, Th17 and Th22 types that implement autoimmune inflammation in the psoriatic plaque. The aim of our work was to study the differences in the skin microbiota spectrum and the parameters of local immunity in capillary blood taken near the focus of inflammation in patients with autoimmune (psoriasis) and allergic (eczema) diseases compared with the parameters of healthy people. 24 patients with psoriasis (group 1), 20 patients with eczema (group 2) and 20 healthy adults (group 3) were examined. Biological materials were taken, i.e., the smears taken with sterile dry swab to the Amies transport medium with activated carbon, and capillary blood was taken in 2 microvets, 200 µL each) from the foci of inflammation on affected skin from the hands of patients, or from the fingers of healthy people. Inoculations of diagnostic media, microscopy with Gram staining and microbial identification were performed in a microbiological analyzer. Immunophenotyping of 22 subsets of mononuclear cells was performed by four-color staining of capillary blood with erythrocyte lysis and evaluation of subsets by a flow cytometer. Cytokines in blood plasma were determined by multiplex method. The spectrum of hand skin microflora of the group 3 was more diverse in the species composition. In patients with psoriasis and eczema, the coccal flora dominated, with a shift towards pathobionts in the microbiota spectrum. Activation of T and B cells, production of pro-inflammatory cytokines, IL-23/IL-17/IL-22 axis cytokines and cytokines – markers of epithelial cell damage (IL-25 and IL-33), as well as anti-inflammatory factors insufficiency were detected. Differences in changing parameters of the local immune status in patients with autoimmune (psoriasis) and allergic (eczema) diseases were revealed, thus reflecting the distinct features of immunopathogenesis in these diseases.

Metabolic aberrations underlie many chronic diseases, including autoimmune diseases (AUD). Immune metabolism is an area of immunological research that is actively developing and studying the processes of metabolic reprogramming in immune cells. The regulation of the nuclear factor kappa B (NF-κB) activity, which is involved in the coordination of innate and adaptive immunity, inflammatory reactions and other processes, is being actively studied. The studies on immune metabolism and regulation of NF-κB is a promising direction in searching for new therapeutic approaches in the AUD treatment. The aim of the present study was to evaluate the informative value of NF-κB and the activity of intracellular lymphocyte succinate dehydrogenase (SDH) and glycero-3-phosphate dehydrogenase (GPDH) determined in children with immune-dependent disorders. 350 children with autoimmune diseases were examined: 97 patients with IBD, 72 children with relapsing-remitting multiple sclerosis (MS), 83 pediatric patients with psoriasis vulgaris (PS), and 97 children with autoimmune hepatitis (AIH). The comparison group consisted of 100 conditionally healthy children. Activity of mitochondrial dehydrogenases, i.e., SDH and GPDH, was evaluated by immunocytochemical method. The levels of NF-κB translocation (per cent of cells with NF-κB translocation from cytoplasm to cell nucleus) was determined by flow cytometry, with visualization. Statistical evaluation and plotting were carried out using the Statistica 13.0 software. The highest activity of SDH and GPDH was detected in the population of cytotoxic T lymphocytes and T helper cells, and the lowest activity of the enzymes was registered in the population of B lymphocytes, both in children with AUD and in comparison group. In children with AUD, there was a significant decrease in SDH activity in T lymphocytes, cytotoxic T lymphocytes, B lymphocytes and NK cells against the comparison group (p < 0.01). In children with PS, AIH and IBD, a decrease in SDH activity was revealed in Treg and Th17 cells. The most pronounced decrease in GPDH was characteristic of patients with AH (in T cells, cytotoxic T lymphocytes, B cells, NK cells and Tregs against the comparison group). In children with PS, the activity of GPDH was reduced only in Tregs (p < 0.05). For children with multiple sclerosis, a decrease in GPDH was revealed in populations of T lymphocytes, B lymphocytes and activated T helpers (p < 0.01). In the group of patients with IBD, there were no significant differences in the activity of GPDG relative to the comparison group. A significant increase in the level of NF-κB translocation in T helpers was revealed in all children with AUD relative to the comparison group. In children with AIH and PS, a significant increase in the level of NF-κB translocation was revealed in Treg, Thact and Th17 cells, in children with MS it was found in Treg cells, in patients with IBD, it was registered in Thact against the comparison group (p < 0.05). An inverse correlation was found between the levels of NF-κB translocation in lymphocyte populations, and activity of mitochondrial dehydrogenases in the lymphocytes. The most significant dependencies are characteristic of NK cells and T cell populations, and these correlations are valid for all groups of AUD patientsh. In the course of in vitro experiments with a drug of metabolic action, a decreased number of cells with NF-κB translocation and an increased SDH activity was observed; the degree of SDH activation depended on the cell population type, the greatest changes were detectable in the population of T lymphocytes (by 61%), in B lymphocytes (by 30%), in NK cells (by 19%). The study of the metabolic activity of lymphocytes and the NF-κB signaling pathway allows us to assess the general mechanisms of immunopathological processes in children with autoimmune diseases of various etiologies. As based on the inverse correlation between the level of translocation of NF-κB and the activity of SDH in lymphocytes, one may consider the use of an available immunocytochemical method being an analogue for assessing activity of the NF-κB transcription factor. The studies of immune metabolic correction of immunocompetent cells are a promising direction in the AUD treatment.

Cytokines regulate the activity of hypothalamus – pituitary – adrenal hormonal axis, also affecting thyroid gland and ovaries. However, their exact role in pathogenesis of endocrine diseases is still under study. Altered cytokine secretion in autoimmune thyroid diseases and polycystic ovary syndrome is well known. Meanwhile, the main immunological predictors of severe prognosis in endocrinopathies and biomarkers for administration of targeted immunotherapy have not yet been determined. Therefore, our objective was to study the relationships between the cytokines and hormones in pathogenesis of autoimmune and non-autoimmune endocrinopathies, i.e., autoimmune thyroiditis (AIT), Graves’ disease (GD), nodular and multinodular goiter, polycystic ovary syndrome (PCOS).

101 patients with GD and 105 patients with AIT, 110 patients with PCOS were examined; 51 patients with nodular and multinodular euthyroid goiter and 50 healthy individuals; their average age was 34.5±2.9 years old. The study was approved by the Interdisciplinary Ethics Committee of the Pacific State Medical University. Clinical examination included ultrasonography. ELISA technique was applied for determination of thyroid and sex hormones, TSH, autoantibodies to TPO, TSH receptor, cytokines in blood serum, in follicular fluid samples, cell culture media (primary cultures of cumulus cells). Genetic studies were carried out with PCR. Morphological verification was performed by inverted microscopy CKX41 (Olympus) with a phase contrast system, camera AxioCam5 (Carl Zeiss) with software Zen 2, Blue Edition.

IL-6 and TNFα in the blood serum of patients with PCOS were most significantly increased, Sharply decreased IFNγ/IL-10 ratio in blood serum and cumulus cell culture was found in PCOS when compared with controls. Significant changes in the content of thyroid hormones in GD and TSH in AIT have been proven to affect the hyperproduction of pro- and anti-inflammatory cytokines. Before treatrment, the direct or reverse corelations were found between the levels of cytokines and thyroid hormones in GD, and TSH in AIT. Subsequently, upon correction of hormonal disorders, these associations became weaker, or faded away. In patients with nodular and multinodular euthyroid goiter, only the IFNγ level was significantly increased, being twice as high as in the control group.

A significant imbalance in the ratio of Th1/Th2 marker cytokines and their hyperproduction in autoimmune thyroid diseases made it possible to characterize the cellular response system in autoimmune thyroid disorders as highly active and directly associated with thyroid dysfunction, performing its effector function under the impaired immunoregulation. The revealed changes in pro-inflammatory cytokines in polycystic ovary syndrome represent new immunological markers of fertility, which may be a promising target for pathogenetic immunotherapy.

Rheumatoid arthritis (RA) is a multifactorial autoimmune rheumatic disease of unknown etiology characterized by chronic erosive arthritis. The protein osteoprotegerin (OPG) is a member of bone tissue homeostasis (RANK/RANKL/OPG) which is responsible for the regulation of osteoclast differentiation and osteolysis. The altered binding of RANKL and OPG is one of the causes of many diseases with increased production of pro-inflammatory cytokines, including rheumatoid arthritis. Polymorphic variants of the genes that control protective reactions could affect the level of production for encoded proteins and, thus, changing the course of immune response in RA. G1181C SNP in osteoprotegerin gene leads to disruption of its transcriptional activity and conformation of the protein itself, which can lead to an imbalance of pro- and anti-inflammatory cytokines. We analyzed the relationship between the TNFRSF11B gene polymorphism at the 1181 G > C position and the risk of developing RA in the Bashkir population. The analysis was based on a molecular genetic study of single nucleotide polymorphism in groups of patients with rheumatoid arthritis and conditionally healthy individuals of the Bashkir population of the Chelyabinsk Region. Statistical evaluation was carried out using standard criteria generally accepted in immunogenetics. The 1181*GC polymorphism of the osteoprotegerin gene is characterized by interpopulation differences, which confirms the importance of using an ethnically identical comparison group to assess the association with a predisposition to multifactorial pathology. We showed that the carriage of genotype 1181 G/C was increased in the group of Bashkirs with rheumatoid arthritis. This genotype could be considered a biomarker of susceptibility to RA. No differences in the SNP allelic frequencies and genotypes were found among women with RA. Our research is a part of comprehensive assessment of the interaction of cytokines and their receptors from the TNFα superfamily as an immunogenetic component of RA genesis.

Many studies have shown that the level of cell-free DNA (cfDNA) in blood of patients with oncological diseases, sepsis, systemic lupus erythematosus, and some rheumatic diseases significantly exceeds the value of similar index in healthy donors and is closely related to the clinical features of the disease. Systemic inflammatory response is among the most frequent pathophysiological processes along with markedly changed levels of cfDNA in blood plasma. The levels of cfDNA in blood plasma of patients with RA are shown to be closely associated with a shifted balance of helpers to the Th1-side. It is an adequate intensity index of inflammatory processes and effectiveness of therapy. At the same time, there only limited number of works concerning changes in cfDNA levels in pathological processes with predominance of Th2 lymphocytes. According to generally accepted concept, the pathogenesis of bronchial asthma is of distinct interest, being critically dependent on the production of specific antibodies controlled by activated Th2 lymphocytes. The aim of this work was to study the level of cfDNA in blood and compare its changes with intensity of NETs and inflammation in patients with asthma. The study included 20 patients with asthma, who underwent hospital treatment at the Department Allergology (Clinic of Immunopathology, RIFCI, Novosibirsk), and 10 conditionally healthy donors. We have shown that, upon admission to the clinic, the level of cfDNA in patients with asthma was significantly reduced against the control group of healthy donors. After a course of therapy, the average level of cfDNA in patients’ plasma was increased and did not differ statistically significantly from this index in controls. The data obtained for other parameters indicate that the patients with asthma did not reveal any signs of pronounced systemic inflammatory response. One should suggest that the observed changes in the level of cfDNA in blood plasma in bronchial asthma are not caused by chronic inflammatory process in lungs of these patients, but they are determined by some other pathophysiological mechanisms. It has been shown that the level of in vitro stimulated NETs in patients with asthma is higher than in healthy donors, thus being consistent with current opinions on the role of neutrophils in pathogenesis of asthma.

Immunological characteristics of airway inflammation in asthma patients with sensitization to various fungal allergens are not well understood, and the search for new markers is necessary to establish future targets for targeted therapy. The purpose of our study was to assess the levels of eosinophilic inflammation markers of the respiratory tract in patients with severe asthma and allergic bronchopulmonary aspergillosis, depending on the spectrum of fungal sensitization.

The study included 31 patients with severe asthma with fungal sensitization and 29 patients with allergic bronchopulmonary aspergillosis (ABPA). The levels of total, specific IgE to fungal allergens and periostin in blood serum were determined by enzyme immunoassay. The study of basophil activation was performed by flow cytometry.

The patients with severe asthma and sensitization to Aspergillus spp. and Alternaria spp. had significantly higher levels of eosinophils, periostin, and stimulation index to Alternaria spp. in the basophil activation test when compared with the group of asthma patients with sensitization to Aspergillus spp. only. In patients with ABPA with combined sensitization, we have found significantly higher levels of eosinophils, periostin, and stimulation index to Alternaria spp.

The finding of pronounced eosinophilic type of inflammation in patients with asthma and combined sensitization to thermotolerant and thermolabile fungi may result from aggressive effect of fungal allergens on the barrier functions of bronchial epithelium, which should be taken into account when choosing therapeutic strategy and administration of immunobiological therapy.

A couple of decades ago, the diagnostic search for the cause of blood eosinophilia concerned, mainly, the three major allergic diseases (bronchial asthma, allergic rhinitis, atopic dermatitis), or parasitic invasion. In recent years, more and more complex clinical syndromes, from reactive (secondary) eosinophilia to eosinophilic leukemia are increasingly considered by the doctors of different specialties. The aim of our work was to present a clinical case of secondary eosinophilia in a child with minimal clinical manifestations. Highlights: Sometimes blood eosinophilia in a patient is an “unexpected finding” for a physician, especially if the complaints are scanty, nonspecific, and objective examination does not reveal any significant health abnormalities. In the presented case, secondary (reactive) peripheral blood eosinophilia was diagnosed due to intestinal damage induced by food allergens. This clinical case is of practical interest to physicians, presenting a diagnostic search for the cause of blood eosinophilia, which eventually proved to be a mixed IgE/ non-IgE mediated food allergy manifesting as allergic enterocolitis. The IgE-mediated mechanism of food allergy is evidenced by a high level of IgE and its decrease with administered elimination diet; delayed-type response and low severity of clinical manifestations of food allergy, as well as blood eosinophilia suggest a non-IgE mediated food allergy. In this particular case, the severity of laboratory changes (blood eosinophilia, a significant increase in the levels of specific IgE) was associated with scarce intestinal symptoms. An opposite situation is observed, especially, in young children where the disease manifests with a pronounced clinical pattern of enterocolitis in the absence of laboratory changes.

Acute kidney injury (AKI) is among most dangerous and common complications after oper-heart cardiosurgical operations. Therefore, a search is carried out for biological markers which could timely detect this condition. The article presents dynamics of TGF-β1, TGF-β2, TGF-β3 in blood serum prior and after a coronary artery bypass (CAB). The study included 120 patients with multivascular affection of coronary blood flow , and 50 conventionally healthy persons of similar age. The 1^st group included 50 patients without evidence of AKI, the 2^nd group consisted of 70 patients with AKI. Serum TGF-β1, TGF-β2, TGF-β3 was determined by ELISA technique in the main groups before (1) and after surgery (2) as well as on day 2 after operation (3), on day 7 after surgery (4), and once tested in the control group.. The results were expressed in ng/mL or pg/mL, as median values, upper and lower quartiles. Significance levels were determined by the Wilcoxon criterion. We have revealed dynamic changes of TGF-β levels in serum of the patients before and after CAB. Initially, before operation, we have found normal TGF-β1 levels and low TGF-β3 levels in the both main groups. Meanwhile, increased TGF-β2 levels are found only in the subgroup with subsequent AKI development. The dynamics of TGF-β1 showed a decrease just after surgery and 2 days later, being increased over initial level on day 7, and there were no significant differences for the groups with versus without complications. No dynamic differences were revealed for TGF-β2 in the patients of group 1 after surgery. Meanwhile, the group 2 after CAB displayed higher TGF-β2 values compared with controls during the entire follow-up period, neing, however, higher that in the group on the 2^nd day following surgery. The TGF-β3 levels were increased just after surgery in both groups followed by subsequent decrease in group 1. In the 2^nd group after CAB, the initial deficiency of TGF-β3 was changes in wave-like mode, over 2^nd and 3^rd period of monitoring. It was increased on day 7, becoming higher than in group1 but did not reach reference values. Further studies in the AKI group after CAB which depend on their severity and outcomes may detect new features of TGF-β system in the patients with this disorder.

In recent years, traumatic brain injury (TBI) has been one of the most urgent medical and social problems due to its prevalence, predominantly affecting young people of working age, causing high mortality, disability and economic costs for treatment and subsequent rehabilitation of patients. At present, the role of patients’ immune system in evolving neuroinflammation after traumatic brain injury has been proven. Treg cell populations represent an important factor determining the outcome of the disease due to promoting induction of immunological tolerance, being a significant component of immunoregulation. As a result of our study, we found a decrease in the relative content of Treg within the total lymphocyte pool of peripheral blood, which has the CD3⁺CD4⁺CD25^bright phenotype in patients of the 3^rd and 4^th groups, in comparison with the data from control group. Moreover, a decreased relative content of Tregs (CD4⁺CD25⁺T cells) was revealed which is due to the degree of brain tissue damage and, as a result, their migration to suppress inflammation due to production of anti-inflammatory cytokines (TGF-â, IL-10). The Treg population is heterogeneous, thus prompting us for analysis of the Treg subpopulations profile based on the expression of CD45R0 and CD62L. When assessing subpopulations of regulatory T lymphocytes within CD45Ro and CD62L, significant changes were found only in patients with brain contusion. The changes were revealed within the pool of “naive” T regulatory lymphocytes with the CD3⁺CD4⁺CD25^brightCD39⁺ Treg phenotype, capable of producing a wide range of cytokines specific for Th1, Th2, Th17, in patients with mild, moderate and severe TBI. Meanwhile, the level of highly active CD3⁺CD4⁺CD25^brightCD73⁺ Tregs was significantly reduced in patients with moderate and severe TBI. These changes indicate an imbalance in immunoregulatory processes resulting from extensive damage to brain tissues.

Autism Spectrum Disorder (ASD) is a complex neurodevelopmental disorder with unknown etiology, high clinical heterogeneity and marked aberrations of the immune system. Evidence for an association between immune dysfunction and behavioral traits highlights the need for a study of the immune cell functional activity in order to search for pathogenesis mechanisms and potential targets for therapy at ASD. The purpose: to determine the expression levels of IL-1β, IL-2, IL-4, IL-6, IL-10, IL-18, IFNγ and TNFα in peripheral blood leukocytes of children with mild and severe ASD. The study included 81 children with ASD (77.8% boys) and 45 children with typical neurodevelopment (TDC, 71.1% boys). According to the Childhood Autism Rating Scale, 51 children (63.0%) had mild autistic symptoms (CARS score 32.0±1,5) and 30 children had severe ASD symptoms (CARS score 39.0±3,4). Cytokines expression in leukocytes was determined by quantitative PCR with SYBRGreen. The data were transformed using Box–Cox transformation. The differences between groups were assessed by one-way ANOVA and Dunn’s test for multiple comparisons. In leukocytes of children with ASD, regardless of the severity, the expression of IL-1β, IL-18 and IL-2, was significantly reduced compared to TDC. Moreover, in children with mild ASD, low expression of TNFα, compared with TDC was found. In children with severe ASD, the expression of the main cytokine of Th1 – IFNγ, was significantly increased, without an increased expression of an important cytokine of Treg – IL-10. Activation of the Th1 adaptive immune response without compensation by cytokines of Treg, the number of which is reduced in ASD, can lead to increased inflammation, even in the central nervous system, and correlates with the severity of ASD clinical symptoms. Despite extensive immunological evidence suggesting immune system dysregulation, further research is required to clarify the relationship between immune system cell function and ASD pathology.

The disease caused by the highly contagious SARS-CoV-2 virus – “novel coronavirus infection (COVID-19)” – had killed more than 6.5 million people at the end of December 2022. The severity of the manifestation of the infectious process varies from asymptomatic forms to rapid progression to life-threatening conditions requiring emergency measures. One of the factors, the severity of which affects the outcome of the disease, is lymphopenia, the cause of which may be a violation of lymphopoiesis. The identification of laboratory markers of a high risk of mortality in patients with COVID-19 plays an important role in improving patient care algorithms and increasing their survival. Levels of TREC and KREC molecules in peripheral blood, respectively, can serve as molecular markers of the severity of T and B lymphopenias. The aim of our work was a comparative analysis of the levels of TREC and KREC molecules in the peripheral blood of surviving and deceased patients with COVID-19. The material was whole blood samples obtained from 1745 people, including: 1028 patients diagnosed with novel coronavirus infection (COVID-19) (ICD-10 code – U07.1), of which 937 patients recovered and 91 died; 717 apparently healthy individuals (control group). The levels of TREC and KREC molecules were assessed by quantitative multiplex Real-time PCR using the TREC/KREC-AMP PS reagent kit (Federal Scientific Research Institute Pasteur, St. Petersburg). Statistically significant differences in the levels of KREC and TREC molecules between the control group and patients, both surviving and deceased, were established. A significant decrease in median concentrations of KREC molecules was shown in patients with a lethal outcome compared with survivors (p = 0.0019, 95% CI). Among the deceased patients, in 63.7% of cases, the levels of TREC or KREC molecules were reduced relative to the corresponding age norms. Of these, in 20.9% of cases, both analytes were reduced in patients. When assessing the diagnostic significance of the levels of the analytes under study for predicting the outcome of the disease, the area under the AUC curve for KREC was 0.63±0.029, which indicates the average strength of the prognostic model of the patient's death depending on the level of KREC in the blood. The constructed model is statistically significant (p = 0.002). Monitoring laboratory parameters of patients with COVID-19, including those who died, allows you to determine the prognostic factors that are most significant for assessing the outcome of the disease. Based on the assessment of the KREC level, a predictive model with high specificity reflects the risk of death in patients with COVID-19. Thus, the quantitative determination of the level of KREC molecules in the peripheral blood can be attributed to the methods of preventive personalized diagnostics aimed at improving the survival of patients.

There are increasing data concerning changes in hematological (clinical) and biochemical blood tests in patients with COVID-19 infection, which indicate the severity of the manifestations of the infectious process. Coagulopathy often correlates with the severity of COVID-19 disease and the risk of death. In this regard, prediction of developing coagulopathy and its prevention remain quite relevant. The aim of our study was to identify differences in the content of platelets and D-dimer in patients with COVID-19. The study included cohorts of patients vaccinated against SARS-CoV-2, and those not immunized against this infection.

A prospective, randomized, observational study of the patients’ response was performed in cohorts of 588/52.2% vaccinated (vaccinated) and 588/52.2% non-immunized (non-vaccinated) patients with diagnosed COVID-19 over the period from 23.06.2021 to 01.05.2022. The levels of blood platelets and D-dimer, as well as clinical outcomes of the disease in patients with COVID-19, were studied in dynamics on days 1-2, 5-6 and 10-12 of hospitalization.

Upon admission, the normal value of the blood platelet counts did not differ between the compared groups, being 206.58 × 10⁹ in vaccinated group and 204.85 × 10⁹ in the unvaccinated group, respectively. a moderate increase in the concentration of D-dimer was noted in both groups upon admission, i.e., 2838.60 ng/mL in the group of vaccinated patients and 3242.08 ng/mL among the unvaccinated patients. In the course of the study, we have shown that the dynamics of D-dimer index in vaccinated versus non-immunized persons was similar according to the days of disease, showing an increase from the first day and a trend towards an higher values, starting from 5-6 days. At the same time, the dynamics in the vaccinated patients was somewhat less favorable than that of the non-immunized subjects. In the patients who were not immunized throughout the entire observation period, the platelet count exceeds the levels found in vaccinated subjects, thus suggesting higher risk of thrombosis and cytokine storm.

The data obtained show that the dynamics of D-dimer and platelet counts in vaccinated and non-immunized people is similar on appropriate terms of the illness. However, the changes are more pronounced in vaccinated cohort, but it does not indicate a greater risk of adverse outcomes.

SARS-CoV-2 virus can induce immune system disorders in post-COVID patients, which may persist for an extended period beyond the acute phase of the disease. Therefore, the search for immunocorrection approaches to address the detected disorders is a significant challenge in clinical immunology. This study aimed to investigate the impact of a synthetic peptide derived from the active center of GM-CSF on the immune system of patients with post-COVID immunopathological syndrome. A total of 21 patients who previously suffered with SARS-CoV-2 infection were included in the study. Flow cytometry was used to analyze various immune cell populations, including panleukocyte markers for gated lymphocytes (CD45⁺ and CD46⁺), T lymphocytes (CD3⁺), helper inducers (CD3⁺, CD4⁺), cytotoxic T lymphocytes (CD3⁺, CD8⁺), TNK cells (CD3⁺, CD56⁺), natural killer cells (CD3^-, CD56⁺), B lymphocytes (CD3^-, CD19⁺), activated helper cells (CD3⁺, CD4⁺, CD25⁺), and activated T lymphocytes (CD3⁺, HLA-DR). Moreover, IgA, IgM, IgG antibodies specific to SARS-CoV-2, phagocytosis and NBT activity of neutrophils, and complement fragments C1q, C3a, and C5a were assessed. The results demonstrated that topical application of the synthetic peptide derived from the active center of GM-CSF (Acegram-spray) upon mucous membranes significantly influenced the functional bactericidal activity of neutrophils (NBT-activity), increased the percentage of T helper cells, and elevated the C3a complement fragment. These findings indicate that the synthetic peptide primarily affects innate immunity factors. However, no significant differences were observed in other immune system parameters. Therefore, the development of therapeutic approaches for post-COVID patients with impaired immune systems may require a search for additional immunomodulators that target T, B, and NK cells. Further research is needed to explore the effects of various immunomodulators on the immune system of post-COVID patients.

The mucosal immunity performs an important function in prevention of respiratory infections including COVID-19. The search for approaches to activate the synthesis of post-infectious antibodies by correcting the factors of innate and adaptive immunity at mucous membranes of respiratory tract in patients with infection caused by the new coronavirus may be relevant for the treatment of patients with COVID-19. The aim of our study was to assess the concentrations of sIgA in the upper respiratory tract in patients with a confirmed diagnosis of Coronavirus infection caused by the COVID-19, and to evaluate the effect of an immunostimulating drug of bacterial origin upon the sIgA secretion.

The patients were divided into two groups: group 1 (n = 45), received basic therapy; group 2 (n = 33), in addition to basic therapy, received the bacterial vaccine Immunovac-VP-4 according to a combined scheme. The biomaterial sampling was carried out by scraping of epithelial cells from the nasal mucosa, pharyngeal scraping and salivary gland secretion on days 1, 14 and 30 of the study. sIgA levels in all biological fluids were studied using ELISA technique (JSC Vector-Best, Russia).

14 days after the start of observation, the dynamics of sIgA levels in nasal scrapings in group 1showed a significant decrease relative to the baseline values (p = 0.02), whereas the level of sIgA remained unchanged during the specified period (p = 0.07) in the group of patients receiving, along with basic therapy, additional Immunovac-VP-4 treatment. The dynamics of sIgA level in pharyngeal scrapings in the group of patients receiving only basic therapy did not change throughout the study period. Menwhile, the group of patients receiving basic therapy supplemented with Immunovac-VP-4 showed a significant increase in sIgA levels by the 30^th day of follow-up over the baseline values (p = 0.02). The level of sIgA in salivary gland secretions did not differ significantly between the study groups during the entire follow-up period.

The results of our study showed that, in order to assess the state of mucosal immunity in patients with COVID-19, one may recommend determination of sIgA in nasal secretions. The Immunovac-VP-4 prescribed in complex therapy is accompanied by an increase in the sIgA levels at the mucous surfaces of the respiratory tract.

Acute destructive pneumonia (ADP) is a severe purulent and septic infectious disorder of childhood, characterized by a high level of morbidity and associated with imbalance of the immune system (IS). Hence, there is an obvious need to study the immunopathogenesis of this disease in order to develop new therapeutic strategy aimed at eliminating the pathogen, detoxifying the body, relieving respiratory failure and correcting functional immune deficiency. Our aim was to perform a clinical and immunological study in order to evaluate efficiency of immunomodulatory therapy using a medical drug with hexapeptide as an active substance. This drug was included into the complex postoperative treatment of children with acute destructive pneumonia. Clinical and immunological examination of 15 children 2-5 years old with ADP was performed before (study group 1 – SG1) and after (study group 1a – SG1a) combined postsurgical treatment including immunomodulatory therapy with a Hexapeptide-based pharmaceutical (HP, Arginyl-alpha-Aspartyl-Lysyl-Valyl-Tyrosyl-Arginine). Comparison group (CG) included twenty healthy children. The contents of T and B lymphocytes, natural killer cells (NK) were measured by means of flow cytometry (CYTOMICS FC 500, USA). Serum levels of IgA, IgM, IgG (ELISA), phagocytic and microbicidal activity of neutrophil granulocytes (NG) were also evaluated. Prior to the treatment in SG1 patients, a decreased number of CD3⁺CD19^-T lymphocytes, CD3⁺CD8⁺ T_CTL lymphocytes was revealed along with significant decrease in the contents of CD3^-CD16⁺CD56⁺ NK (p_1-3 < 0.05). It was found that in children with ADP, the IgG level did not differ from indices of control group, with a decrease of IgM (p_{1, 2} > 0.05), and increased level of IgA (p < 0.05). We have also found a deficiency of NG effector functions, i.e., insufficiency of active phagocytic NG with impaired capture and killing of bacterial antigen. Assays of NADPH-oxidase activity showed lacking response to both inflammation and additional induction by S. aureus. After complex treatment including immunomodulatory therapy in the SG1a group, we revealed a recovery in CD3⁺CD19^-T lymphocyte contents, CD3⁺CD8⁺T_CTL lymphocytes, CD3^-CD16⁺CD56⁺ NK (p_1-3 < 0.05). The trends towards normalization of IgA, IgM, improved NG effector functions (killing ability) were revealed due to activation of NADPH-oxidases. The restoration of immunological parameters in ADP was associated with earlier recovery from the purulent-destructive process in lungs, absence of postoperative complications including the prevention of septic process. The clinical and immunological effects of the immunomodulatory therapy program with HP-based pharmaceutical preparations suggest its potential usage during postoperative period in immunocompromised children with ADP.

Negative impact of S. aureus, seems to be a sufficient condition for the spread of the infectious process in the bone in acute osteomyelitis (AOM) due to its altered elimination caused by dysfunction of the immune system (IS), in particular, of neutrophilic granulocytes (NG). Correction of NG dysfunction in AOM under the influence of immunotropic substances and cytokines via modulation of the NG phenotypic subsets is of sufficient interest. Our aim was to evaluate the in vitro effects of recombinant IFNá2b on the number and phenotype of CD66b⁺CD16⁺CD33⁺HLA-DR^-, CD66b⁺CD16⁺CD33⁺HLA-DR⁺ subsets and on phagocytic function of neutrophilic granulocytes in acute osteomyelitis in children.

The study of peripheral blood (PB) samples from children aged 8-15 years was carried out as follows: patients with АOM (n = 24) comprised study group 1 (SG1), healthy children (n = 13) were included into comparison group (CG). PB samples of children with AOM were incubated with recIFNá2b (50 IU/µL, 60 min, 37 °C.) in the study group 1a (SG1a). Before and after incubation with recIFNá2b, the number of NG subsets CD66b⁺CD16⁺CD33⁺HLA-DR^-, CD66b⁺CD16⁺CD33⁺HLA-DR⁺ and the density values of receptor expression by fluorescence intensity (MFI) were also determined (FC 500, Beckman Coulter, США). Phagocytic activity of NCs was evaluated as the contents of actively phagocytic NCs (%PhAN), volume of the engulfed S. aureus (strain 209) by assessing their phagocytic number (PhN), phagocytic index (PhI). Bacterial killing was determined as the percentages of microbe digestion (%D), digestion index (DI).

The cells from AOM patients revealed a subset expressing the HLA-DR receptor – СD66b⁺CD16⁺CD33⁺HLA-DR⁺NG, which is absent in the PB of CG children. The cells with primed phenotype exhibited an increased expression density of activation receptors CD16 and CD66b. Incubation of PB in AOM with recIFNá2b led to an increased proportion of CD66b⁺CD16⁺CD33⁺HLA-DR⁺ NG subset which showed active phagocytosis and improved digestion processes. The present study shows the emergence of activated subset of “long-lived” CD66b⁺CD16⁺CD33⁺HLA-DR⁺ NCs in children with AOM. This subpopulation has APC features, by presenting AG to T lymphocytes, with preserved effector properties. In an in vitro experimental system, a positive effect of recIFNá2b was demonstrated, leading to an increased number of NGs of the CD66b⁺CD16⁺CD33⁺HLA-DR⁺ subset and recovery of S. aureus phagocytosis by NGs, thus being promising in the future for development of new approaches to optimization of complex therapy in the postoperative period of AOM treatment, prevention of complications and the opportunity to alleviate the disorders in the immune system.

The COVID-19 pandemic, caused by SARS-CoV-2, remains one of the actual medico-social issues of today's world. Novel coronavirus infection officially listed in dangerous infections. When an ingress of coronavirus infection in the background of intensive production of inflammatory inducers comes with decreased levels of type I interferon that cause loss of protective abilities of the body against the background of the destruction of its own tissues. Herd immunity development via vaccination increases the proportion of people with protective immunity against SARS-CoV-2 is an important factor in stopping the spread of the infection. However, during the first days after vaccination, patients remain susceptible to possible infection. A promising prophylactic agent is interferon-containing drugs widely used in Russia and CIS countries for the prevention and treatment of viral infectious diseases, in particular SARS and influenza. Our study showed that intranasal recombinant IFNα-2b has clinical and immunological efficacy after two courses of the drug (within 5 days after V1 and after V2 vaccination with the EpiVacCorona vaccine).