The processes of adaptation of the human immune system to negative internal and external influences, including stress, infectious, environmental factors, are controlled by the neuroimmunoendocrine complex (NIEC). The normal functioning of the NIEC is the close interaction between the nervous, immune and endocrine systems. The normal NIEC allows the immune system show its adaptive capabilities, remain resistant to various negative influences, while maintaining the ability to repel attacks of various pathogens, neutralize allergic and autoimmune reactions. In case of violations of the normal functioning of the NIEC, adaptive processes in the immune system, the interaction of the immune, nervous and endocrine systems suffer, which leads to persistent violations of the functioning of the immune system and can manifest the formation of diseases of the immune system, including acquired immunodeficiency. We have justified the need to create a new promising scientific direction – adaptive medical immunology, which is based on the study of the features of disorders of the human immune system under acute or chronic exposure to negative internal and external, stressful, infectious, urban factors, resulting in a “breakdown of the adaptive capabilities of the immune system” and, as a result, the diseases exist of the immune system, including acquired immunodeficiency and related immune-dependent diseases. The aim of adaptive medical immunology is to develop an algorithm for complex immunodiagnostics and to create integrative interdisciplinary programs for correction immune system, including immunotherapeutic approaches that affect acquired disorders and restore an adaptive activities of the immune system. The main task is the restoration of an adequate response of the immune system to the existing atypically occurring acute or chronic infectious and inflammatory process, as well as the leveling of disorders of the nervous and endocrine systems with restoration their balanced interaction in the NIEC. Such an integration approach contributes to the restoration of the normal functioning of the immune system, the formation of positive clinical efficacy and the exit of patients into long-term clinical remission. Another task of adaptive medical immunology is the creation of promising innovative diagnostic technologies and, on this basis, the development of new integration programs for targeted restoration of damage to the immune system, as well as the nervous and endocrine systems in immunocompromised patients suffering from various immune-dependent diseases.

Mast Cell Activation Syndrome (MCAS) is a severe relapsing disease requiring inpatient treatment, with clinical pattern including the features of anaphylaxis. The article presents diagnostic criteria aimed for differentiation of MCAS from similar severe conditions as well as discusses local forms of mast cell activation. The consensus group has established distinct criteria for diagnosing MCAS. The agreed criteria include episodic (recurrent) occurrence of typical systemic symptoms caused by release of mast cell mediators and involve, at least, two organs; an increase in serum tryptase level by, at least, 20% over individual baseline tryptase plus  2 ng/mL tryptase during 3-4 hours of the pathological reaction; a positive response to drugs that either target mast cells mediators, or their effects. In principle, the classification of MCAS is based on its etiology being subdivided into primary (clonal) MCAS, secondary MCAS, and idiopathic MCAS. The primary MCAS is determined by clonal expansion of mast cells and is considered systemic mastocytosis. In secondary MCAS, normal mast cells are activated by the known triggers, e.g., IgE. If neither clonal expansion nor a trigger for mast cells activation are identified, the condition is defined as idiopathic MCAS.

The new COVID-19 infection has attracted particular interest in MCAS, since the severe course of COVID-19 was thought to develop due to latent MCAS, but the criteria for MCAS in these patients were not reproduced. In the presence of local symptoms, such as urticaria, or in cases of single-organ involvement, e.g., isolated gastrointestinal symptoms, and suspected mast cell activation being basic to pathogenesis, the term mast cell activation disorder was introduced. Moreover, the article discusses several different mediators that are proposed as markers in the diagnosis of MCAS.

However, over-diagnosis of MCAS entails the risk of missing the underlying pathology, which is not associated with MCAS, and requires differential diagnosis with a number of diseases. In the absence of severe attacks (with hypotension and shock), the likelihood of MCAS is generally very low. Of course, the patients with mastocytosis and/or confirmed IgE-dependent allergy are at higher risk of developing MCAS, but a   key diagnostic marker is an event-related increase in mast cells tryptase from baseline determined over the asymptomatic period. The diagnosis of MCAS is highly likely if the tryptase level rises above a certain threshold (20% of baseline plus 2 ng/mL).

The review article contains data from literature which concern the role of Toll-like receptors (TLRs), immune sensors that play a key role in the systemic response to both bacterial and viral infections, e.g., in pathogenesis of a new coronavirus infection (COVID-19, SARS-CoV-2 infection). With advent of COVID-19, which has reached the scale of a pandemic, the interest in studying predictive factors for the severity of the infectious process has acquired a new cycle. The previous epidemics caused by severe acute respiratory syndrome virus (SARS-CoV), as well as the Middle East respiratory syndrome coronavirus (MERS-CoV), helped us to understand the degree of immune response in these conditions, as well as to suggest medical approaches to the pathogens of this family, i.e., which measures should be taken, and what long-term forecasts may be encountered for the SARS-CoV-2 outbreaks. Each of the 10 human TLRs recognizes a specific structure within a bacterial / viral or fungal pathogen. The effect on TLR activates the inflammatory signaling cascade via mediators, i.e., intracellular TIR domains mediated by adapter proteins. These reactions lead to the production of the most important antiviral response substances. The factors that lead to reduced / increased expression of TLR genes include gene polymorphisms which control the functioning of the immune system in some ways, thus causing a reduced, or hyperinflammatory response to an infectious agent. Genetic heterogeneity is likely to explain, at least partially, the wide range of clinical manifestations of COVID-19 infection in general population. Therefore, there is an increased interest in studies of these receptors, the degree of their expression throughout the infectious process, the polymorphisms of the TLR-encoding genes, and, consequently, the opportunity of using clinical and laboratory tests for their qualitative and quantitative assessment, as well as selection and prospects of further treatment in each personal case.

Disorders of immune homeostasis represent the key pathogenetic link of COVID-19, which often manifests as a hyperimmune response to the pathogen, leading to severe uncontrolled inflammation in lungs, followed by complications and death. Accordingly, a certain therapeutic potential is provided by different pharmacological drugs with distinct mechanisms of action, This class of drugs should, however, act in common direction by suppressing the immune response, thus being often classified as immunosuppressants (IS). Of them, the most promising are immunobiological preparations, which include monoclonal antibodies, as well as purinergic regulatory agents. There are several attempts to use the “classical” IS by a certain way, e.g., cytostatics and calcineurin inhibitors which found clinical application in transplantology and oncology. However, their usage for treatment of uncontrolled inflammation of respiratory tract was abandoned by the end of XX century. Meanwhile, the aerosol route of drug administration optimizes treatment, both in terms of their effectiveness, and the reduction of side effects thus promoting usage of IS for treatment of uncontrolled airway inflammation. Previously, the analysis of therapeutic opportunities for some IS delivered as aerosols to the lungs in COVID-19 therapy was not carried out, thus bein the purpose of our work. Methodological analysis was carried out using various databases of biomedical scientific information, including Index Medicus, PubMed, Embase, Cohrane Clinical Trials gov registry and patent databases.

The efficiency of the impact of various IS subgroups in COVID-19, including their administration by inhalations into the respiratory ways, was assessed. The role of regulatory T cells considered the central regulator of immune response, in pathogenesis of COVID-19 was considered, and their therapeutic potential was characterized, dependent on phase and severity of the disease as well as drug dose dependence. Methods and approaches to the use of IP, advantages and disadvantages are discussed. The expediency and future prospects of their application are considered.

One may conclude that the effectiveness of cytostatics and calcineurin inhibitors in the treatment of airway inflammation in COVID-19 remains unconfirmed and seems to be unpromising. Meanwhile, biological preparations, including monoclonal antibodies and purinergic regulatory agents, offer great promise in this respect.

It is well known that ischemia and hypoxia in the tumor microenvironment promote tumor progression. Оxygen deficiency in tumor microenvironment polarizes cancer cell metabolism from oxidative phosphorylation to the aerobic mode (Warburg effect) and anaerobic glycolysis. This altered carbohydrate metabolism is characterized by low energy efficiency and excessive glucose consumption. Under hypoxic conditions, the antioxidant protection of malignant cells becomes weaker, thus causing a sufficient increase of their susceptibility to direct toxic effects of reactive oxygen species (ROS). In clinical practice, oxygen saturation of tumors is usually achieved by using water-soluble ozone or hyperbaric oxygen. The ROS are shown to be produced by oxidative burst, thus being able to enhance antitumor effects of chemoradiotherapy. The immune cell-derived ROS were shown to directly inhibit tumor growth. In addition, ROS provide additional immune stimulation through the induction of mutagenesis in the tumor cells and production of immunogenic neoantigens. ROS may also enhance antitumor immune defense by inducing synthesis of interferon-γ, tumor necrosis factor-α, IL-2 and IL-6 by immune cells. Moreover, ROS may exert a negative effect on antitumor immunity. In particular, they are able to: (I) favor the recruitment and accumulation of regulatory T cells (Treg) and myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment; (II) promote accumulation of alternatively activated (M2) macrophages and (N2) neutrophils, and, (III) impair presentation of immunogenic antigens (Ag) by dendritic cells. We suggest that the maximal clinical effect of oxygen therapy could be achieved in case of its simultaneous or sequential combination with immunotherapeutic interventions.

The authors conclude that:

–         oxidative stress-induced reactive oxygen species may preferentially damage tumour cells without significantly affecting normal cells;

–         oxygen therapy may potentiate anti-tumour effects of chemoradiotherapy;

–         oxygen therapy could be effectively combined with immunotherapy to achieve maximal anticancer effects with minimal side effects.

Glycodelins, the glycosylated proteins of reproductive tract are characterized by immunomodulatory functions, are of interest because of their role in the development of immune tolerance. Interleukin-17-producing T helpers (Th17) bearing the surface marker CCR6, are a heterogeneous cell population with increased plasticity and functional dichotomy. On the one hand, these cells support antimicrobial and antifungal immunity and microbiota composition; on the other hand, they are involved in the pathogenesis of autoimmune diseases, graft rejection, and pregnancy complications. Despite the scientific interest in glycodelin as an immunomodulator, its direct effects on pro-inflammatory Th17 have  not been studied. Therefore, the aim of our work was to investigate the effect of recombinant human glycodelin on Th17 polarization of naïve human T helper cells cells by assessing surface expression of CCR6, CCR4, and CXCR3  molecules. Naïve T helper cells were polarized for 7 days in vitro to Th17 cells with a TCR activator and cytokines for 7 days, supplemented with glycodelin at concentrations appropriate for the 1^st and 2^nd trimesters of pregnancy. The percentages of CD4⁺CCR6⁺ cell population (Th17 cells), and their CCR4⁺CXCR3^-(Th17/Th22) and CCR4^-CXC3⁺ subpopulations (Th17.1) was then determined. Moreover, the levels of IL-17, IL-2, and other cytokines/chemokines were determined in the culture supernatants of Th17-polarized T helper cells. Treatment with recombinant glycodelin at concentrations equivalent to those in pregnancy (0.2, 2, and 10 μg/mL) did not alter the percentage of CD4⁺CCR6⁺ cells in culture, or their IL-17 production. However,  at a concentration of 10 μg/mL, it caused a decrease in Th17.1 (CCR6⁺CCR4^-CXCR3⁺) percentage in the T helper culture, and increased the production of IL-2. In addition, glycodelin was found to have selective pro-apoptotic activity against Th17.1 if applied at 2 μg/mL. Given the known involvement of these cells in pathological processes, the observed effect of glycodelin could be of interest from a biopharmaceutical perspective. However, the mechanism of the revealed selective effects of this pregnancy protein needs further investigation.

The immunological factors can play an important role as predictive and prognostic biomarkers in oncopathology. Recently, non-conventional innate-like γδT-lymphocytes have received a lot of attention as a promising effector cell population for cancer immunotherapy. This study describes structural and functional subpopulations of γδT lymphocytes involved in antitumor immunity in patients with malignant and benign tumors of the nasal cavity and paranasal sinuses. The aim of the study was to estimate γδT cell subsets composition and functions in patients with neoplasms of nasal cavity and paranasal sinuses in order to characterize cellular immunity in tumor-associated pathological process.

The peripheral venous blood was obtained from 21 patients (13 men and 8 women, average age of 63.0 (56.0-69.0) y. o.) with neoplasms of nasal cavity and paranasal sinuses, and 10 healthy donors. Lymphoid cells phenotype and production of intracellular cytokines were investigated using monoclonal antibodies and flow cytometry, production of extracellular cytokines was measured using enzyme-linked immunosorbent assay kits.

The increase of total γδT cells number in patients with squamous cell carcinoma accompanied by changes in Vγ2⁺/Vγ1⁺T cells ratio in peripheral blood of both patients’ groups with malignant and benign nasal cavity and paranasal sinuses tumors were revealed as compared to healthy donors. The upregulated γδT cell response to phosphoantigen induction in combination with reduced indices of stimulations were shown in the both patients groups but cytokine profile was different, i.e., the elevated IFNγ production has been determined in patients with squamous cell carcinoma. However, in patients with inverted papilloma, redistribution of γδT cell subsets has been associated with IL-17-producing γδT cells. Moreover, the percent of IFNγ⁺γδT lymphocytes did correlate with IFNγ concentration in cell culture supernatants of patients with malignant nasal cavity and paranasal sinuses neoplasms (R = 0.61; p < 0.05).

The revealed data suggest an involvement of γδT lymphocytes in malignant and benign tumor pathogenesis and may provide a fundamental basis for further detection of possible tumor-associated inflammation and malignization predictors.

Maternal adaptation of the immune system aimed at limiting the immune response to fetal antigens is a necessary condition for a successful pregnancy. It involves various mechanisms (Th1/Th2 switching, Treg expansion, induction of anergy and apoptosis of T lymphocytes, development of T cell depletion) that are induced through the ligation of inhibitory receptors. Accordingly,  the expression of inhibitory receptors on  T cells, including PD-1, CTLA-4, and Tim-3 molecules, may reflect the effectiveness of immune adaptation and its impairment in pregnancy pathology. Preeclampsia (PE), the pathogenesis of which is associated with the impairments of immunological tolerance is a major complication of pregnancy. Accordingly, changes in the expression of inhibitory receptors on T cells may be biomarkers of abnormal gestation and potential therapeutic targets. The aim of this work was to study the expression of inhibitory molecules on peripheral blood T cells in women with PE. The study recruited 29 pregnant women with PE and 36 women with uncomplicated pregnancies in the second half of pregnancy. Pregnant women of the study groups were comparable in terms of gestational age, number of pregnancies and parity of childbirth. The control group consisted of 28 fertile women with children. Relative content of CD8⁺PD-1⁺, CD8⁺CTLA-4⁺, CD8⁺TIM-3⁺, CD8⁺PD-1⁺TIM-3⁺, CD4⁺PD-1⁺, CD4⁺CTLA-4⁺, CD4⁺TIM-3⁺, CD4⁺PD-1⁺TIM-3⁺T cells in blood were analyzed  by  flow cytometry. It has been shown that uncomplicated pregnancy is associated with increased expression of PD-1 and Tim-3 T cells, which is manifested by an increase in the relative content of CD4⁺Tim-3⁺, CD8⁺PD-1⁺ and PD-1⁺Tim-3⁺T lymphocytes . In PE, on the contrary, there is a reduction in the expression of PD-1 and Tim-3 by T cells, in particular, a decrease in the proportion of CD4⁺Tim-3⁺ and CD8⁺PD-1⁺ cells; the absence of elevated levels in PD-1⁺Tim-3⁺ cells (compared to uncomplicated gestation) and an increase in CTLA-4⁺ cells within CD4⁺ lymphocytes. Changes in the expression of inhibitory receptors are associated with the severity of PE. A decrease in CD4⁺Tim-3⁺ and CD8⁺PD-1⁺T cells is most typical for patients with moderate PE, and an increase in CD4⁺CTLA-4T cells for pregnant women with severe PE. The relationship between changes in the expression of inhibitory molecules and the onset of PE has also been demonstrated. A distinctive feature of early PE is a decrease in the proportion of CD8⁺CTLA-4⁺ cells and a more pronounced increase in CD4⁺CTLA-4⁺ cells, while late PE is associated with a decrease in CD4⁺PD-1⁺ cells and a more pronounced decrease in CD4⁺Tim-3⁺ cells. The results obtained indicate a changes in the expression of CTLA-4, PD-1 and Tim-3 molecules on circulating T cells in pregnant women with PE and the association of these changes with the severity and the onset of PE manifestation.

The avian influenza virus (AIV) has a great pandemic potential both in animals, and in human population. For effective struggle against this virus, it is important to study the immune response against AIV, and, in particular, the features of T cell immunity. At the period of early immune response, when the activation of adaptive immunity factors has not yet occurred, the natural killers, which have both cytotoxic and immunoregulatory functions, are known to play a key role in the fight against the influenza virus. In this study, we investigated the interaction between natural killers and T helpers in the process of antiviral response. The aim of this study was to study immunoregulatory functions of NK cells in BALB/c mice vaccinated with different doses of avian influenza virus.

We used a strain of mouse-adapted avian influenza virus (AIV) of the H5N2 serotype (A/duck/ Pennsylvania/10218/1984). The determination of the virus titer in the mice lung tissue was performed by infecting 10-day-old chicken embryos (SPF) with lung extracts at various decimal dilutions, followed by the immune agglutination test. Quantitative determination of natural killers and T helpers (Th) in the mice spleen tissue was carried out by flow cytometry. The amount of Th cells in mice spleens increased at low infectious dose (2.5 EID50) which is explained by active clonal expansion of this cell subpopulation. The infectious disease process ended upon complete virus elimination from the lungs. The amounts of Th cells were also increased in a group of mice infected with intermediate infectious dose (25 EID50), however, being accompanied by virus replication in lungs. When the mice were infected with a high dose of the virus (250 EID50), there was a decrease in the Th cells number versus control group of animals, whereas replication of AIV was noted throughout the entire observation period in the lung tissue.

The amounts of helper T cells at high doses of viral infection could be decreased due to activation of NK cells. The activated NK cells may then eliminate exhausted helper T cells. Thus, NK cells immunoregulatory function represents an important immunological factor which keeps balance between the pathogen destruction and excessive inflammation of the lung tissue affected by the avian influenza virus.

The most important role in homeostasis of intestinal immune belongs to the immunoregulatory properties of the microbiota which activates intracellular signaling systems, cytokine expression, production of protective factors and limits inflammatory reactions in the intestine by interacting with the pattern recognition receptors. The outcome of interactions between the microbiota and host cells (development of an inflammatory process or maintenance of intestinal homeostasis) depends on many factors, including a potential ability of intestinal commensals to influence the cytokine network in human body. Due to disturbances of quantitative and qualitative microbiota profile (dysbiosis), the cytokine balance may be changed by the influence of intestinal microsymbionts and their metabolites on immune and epithelial cells of intestines, thus contributing to the development of various human disorders. The aim of this study was to evaluate the immunoregulatory properties of eubiotic and dysbiotic human intestinal microsymbionts by assessing the effects of their cell-free supernatants on cytokine production in the in vitro system. The study was conducted on 49 eubiotic and 77 dysbiotic strains of microorganisms isolated from conditionally healthy patients examined for colon dysbiosis. To assess immunoregulatory properties of intestinal microsymbionts, we studied the effects of cell-free supernatants from bacterial and fungal cultures up on production of proinflammatory (IFNγ, TNFα, IL-17, IL-8, IL-6) and anti-inflammatory (IL-10, IL-1ra) cytokines secreted by mononuclear cells isolated from peripheral blood of healthy persons. The intestinal microbiota was determined by bacteriological methods. Identification of isolated microbial cultures was performed using MALDI TOF MS Microflex LT series (Bruker Daltonics, Germany). The level of cytokines was determined by enzyme immunoassay using commercial test systems (“Cytokine”, Russia). Statistical evaluation included discriminant analysis, classification decision tree and resultant mapping method. The multivariate statistical analysis enabled us to determine the range of the most informative indexes among cytokines and microbial cultures that changing their production in order to assess the state of homeostasis in eubiosis and intestinal dysbiosis. It was found that the supernatants of eubiotic cultures of intestinal symbionts exhibited a pronounced ability to inhibit the level of pro-inflammatory cytokines (IFNγ, IL-8) and to stimulate the secretion of anti-inflammatory cytokine (IL-10), whereas the dysbiotic cultures predominantly induced pro-inflammatory cytokines (IL-17, IFNγ, TNFα). In maintaining a uniform balance between pro- and anti-inflammatory cytokines during eubiosis, both associations of microsymbionts (in descending order of factor loads): Bacteroides spp. > E. coli > Lactobacillus spp.), and monocultures (Bifidobacterium spp. and Lactobacillus spp.) made a significant contribution via IL-10 induction. In cases of intestinal dysbiosis, we found an increased number of associations between microsymbionts inducing secretion of pro-inflammatory cytokines was. The pro-inflammatory profile of dysbiotic cultures was determined by the influence on IFNγ production (ranged in descending order of factor loads) of Bifidobacterium spp. > Enterococcus spp. > E. coli > Lactobacillus spp. associations, as well as S. aureus > Candida spp associations. The secretion of IL-17 was influenced by the monoculture of Clostridium spp., and by association C. acnes > S. aureus > Klebsiella spp. Monocultures of Bifidobacteria and Escherichia exerted effects upon TNFα production. Thus, during eubiotic state, the normobiota maintains a uniform balance of pro- and anti-inflammatory cytokines, and, in presence of intestinal dysbiosis, a shift in the balance of cytokines towards pro-inflammatory ones may occur due to increased levels of their secretion, an expanded spectrum of cytokines from this group, and increased number of single bacteria and associations of microbial cultures affecting their production.

Helicobacter pylori (H. pylori) is a common infection which can lead to gastritis, peptic ulcer disease and gastric-originated malignancies. In this study prevalence of seropositivity of each immunoglobulins against H. pylori and also, their association with sex and age were evaluated in a sample of the ordinary population from Tabriz, Iran.

In this study, 3733 individuals referred to the laboratory for Para clinical tests between 2019 and 2022, participated. Enzyme-linked immunosorbent assay (ELISA) was utilized to detect the quantity of anti-H. pylori Immunoglobulin G (IgG), Immunoglobulin M (IgM), and Immunoglobulin A (IgA). The statistical analysis was conducted using the 20th version of SPSS software.

Out of 3733 participants, 1235 (33.1%) were male and 2498 (66.9%) were female. 57.9% of the participants have positive IgG serology. Also, this index was 0.3% and 11.6% for IgM and IgA, respectively. The mean (SD) age was 40.72 (16.91). There was no significant relationship between gender and IgG and IgA positiveness (p-values = 0.11 and 0.08 respectively). For IgM, serum positiveness was higher in females (0.4% for females and 0.2% for males; p-value: 0.009). The prevalence of positive IgG was increasing significantly (p-value < 0.001). For IgM and IgA there was a significant increase in the number of seropositive individuals with an increase in age (p-value = 0.005; and < 0.001 respectively).

This study reveals that the prevalence of H. pylori in Tabriz is approximately 57% which is in the range estimated to be in Iran, but in comparison to developed countries, it was higher. The prevalence of anti-H. pylori immunoglobulins increases significantly with age.

Psoriasis is a chronic autoimmune skin disease with affected T-cell immunity. The interleukin IL-23/IL-17/IL-22 cytokine axis is a key to immunopathogenesis of psoriasis. Certain role of the IL-36 subfamily is shown in regulation of skin inflammation. Topically applied preparations are used to treat psoriasis. Our aim was to evaluate the treatment-related changes in the cytokine profile of venous and capillary blood collected close to the foci of psoriatic inflammation. Forty patients with psoriasis (mean age, 43.7 years), were examined. Group 1a (20 people) received local treatment with Mometasone, Group 1b (20 people) received topical gel containing an IL-36 receptor antagonist. Twenty healthy people (mean age, 46.6 years) comprised the control group 2. 200-μL aliquots of capillary blood were collected in a microvette with EDTA from the patients’ finger near to the lesion area. Venous blood (3 mL) was taken from the cubital vein to a vacuum tube with EDTA. The concentration of 15 cytokines in blood plasma was tested by the multiplex method (MagPix, BioRad, USA). Clinical effectiveness of therapy was assessed using the PASI and DLQI indexes. Upon completion of treatment (day 14), the PASI and DLQI indices were significantly decreased in both groups. On the 28th day, the PASI index in Group 1a returned to its original level, in group 1b it remained permanently reduced. Before treatment, the levels of all cytokines, except of IL-10, were significantly increased in capillary blood samples of patients with psoriasis compared to Group 2, and the levels of five cytokines were increased in the venous blood. In group 1a, the levels of IL-1, IL-4, IL-6, IL-21, IL-22, IL-23, IL-25, IL-33 were significantly decreased in capillary blood after 14 days, and only IL-17F, IL-21, IL-33 and TNF showed a decrease in the venous blood specimens. On the day +28, the concentrations of almost all cytokines returned to their original level. In Group 1b, on the 14^th day, the levels of IFNγ, IL-1, IL-4, IL-17F, IL-21, IL-22, IL-23, IL-25, IL-33 were significantly decreased in capillary blood compared to altered IFNγ, IL-21, IL-22, IL-23, IL-33 in venous blood. On the 28^th day, their concentration continued to decrease, or the level of these cytokines remained reduced, along with significant decrease of IL-6 in venous samples. Thus, the method for determining cytokine profile in capillary blood from the area of psoriatic lesions may be used for tracing the effects of therapy in psoriatic patients.

Breast tumors show a complex structure and are highly heterogeneous. The study of cytokines, which exert great influence on tumor cells, and microRNAs, which, along with their influence on the proliferation and migration of neoplastic cells, may affect the work of cytokines, will contribute to a deeper understanding of pathological processes occurring in breast cancer. The aim of our work was to analyze the relationship of cytokine production with expression of miR-181a and miR-25in patients with invasive breast carcinoma of a non-specific type (IBC NST) with various molecular subtypes.

Patients with IBC NST were divided into five subgroups according to the molecular genetics subtype of the tumor classified by immunohistochemical analysis of estrogen receptor (ER), progesterone (PR), epidermal growth factor 2 (HER2) and proliferation marker Ki-67. Using enzyme immunoassay, the concentration of 14 cytokines was determined in the supernatants of immunocompetent blood cells and tumors: IL-2, IL-6, IL-8, IL-10, IL-17, IL-18, IL-1β, IL-1ra, TNFα, IFNγ, G-CSF, GM-CSF, VEGF and MCP-1. The expression of miR-181a and miR-25 microRNAs isolated from the patients’ blood serum was evaluated using digital droplet polymerase chain reaction (ddPCR).

In the luminal A subtype, cytokine concentrations and expression of miR-181a and miR-25 are significantly lower compared to other subtypes. Patients with the luminal B HER2-negative subtype were characterized by significantly increased expression of both studied microRNAs, especially when compared with the luminal A subtype. At the same time, patients with a triple negative molecular subtype, on the contrary, were characterized by high concentrations of cytokines in the supernatants of tumor samples and blood cells compared to   other subtypes. In the general group of patients with IBC NST, direct correlations were found between the expression of both studied microRNAs and the concentration of vascular endothelial growth factor (VEGF) in the supernatant of tumor samples, which may presume mutual interactions existing between miR-181a and miR-25, and the process of angiogenesis in the tumor.

The levels of cytokines in blood supernatants and tumors in invasive breast carcinoma may vary, depending on distinct molecular subtypes of the tumor. Moreover, they also have direct links with the levels of miR-181a and miR-25 in blood serum. Particularly noteworthy were the results of measuring the cytokines and microRNAs concentrations in luminal A, luminal B HER2-negative and triple negative molecular subtypes.

The development of COVID-19 is accompanied by involvement of various cytokines in pathological process. Their change depends on age, concomitant pathology and some other factors that have not been sufficiently studied in elderly patients with coronary heart disease (CHD). The content of cytokines in blood plasma of patients aged 60-74 in the early period of recovery from COVID-19 also remains unknown. The aim of our study was to determine the content of systemic cytokines in elderly patients with coronary heart disease at the early stages of recovery after COVID-19. The patients aged 60-74 with CHD who had COVID-19 of moderate severity grade (n = 40) made up the main group. in which the study of The cytokine levels were studied in blood plasma 3 to 4 weeks after recovery. The control group consisted of 38 elderly patients with coronary heart disease and negative tests for COVID-19. of cytokines in both groups were determined by means of flow cytometry with “Becton Dickinson FACS Canto 2 (USA)” machine using appropriate reagent sets. We have found that, in elderly patients with CHD at 3-4 weeks after recovery from COVID-19, if compared with elderly patients with CHD without COVID-19, the content of IL-6 in blood plasma was increased to higher degree and at statistically significant difference, up to 32.9±2.3 pg/mL versus 6.5±0.7 pg/mL in the control group (p < 0.001). Excessive content in the main group was also detected for IL-17 in blood plasma which was 25.4±1.9 pg/mL at the early post-COVID-19 period, whereas it was 7.8±0.7 pg/mL (p < 0.001) in the age-matched patients with CHD only. TNFα and IFNγ levels were elevated among 60-74-year-old patients with CHD and COVID-19 at 3-4 weeks after recovery, being 128.6±2.7 pg/mL and 57.6±2.8 pg/mL, respectively.  In control group, the concentration of these cytokines was significantly lower, i.e., 56.3±2.2 pg/mL and 25.9±1.7 pg/mL, respectively (p < 0.001). The changes in contents of other cytokines studied seemed less pronounced, and the level of IL-4 was not significantly different between the both groups. Hence, in elderly patients with CHD and COVID-19 at 3-4 weeks after recovery, IL-6, IL-17, TNFα and IFNγ are the most elevated cytokines at the systemic level.

Timely diagnosis and prevention of cardiovascular diseases is based on markers that detect changes in the athlete’s body at an early stage of disease. To implement this task, it is important to use novel laboratory techniques. We have carried out a comparative determination of immunological markers, specific antibodies to angiotensin, bradykinin, histamine, dopamine, serotonin and functional indicators of the cardiovascular system in athletes of various qualifications. The object of study included athletes of the Russian national teams who underwent an in-depth examination (IME) as part of medical survey. The participants were divided into groups depending on the sport arts and qualifications. Representatives of the group “Cyclic sports” included athletes without a category of 30 people and 29 qualified athletes (1^st step, Candidate Master, Master of Sports, etc.). The “Combat sports” group consisted of 32 people without a category and 31 athletes with qualifications similar to those indicated above. Athletes of “Speed-strength” sports are represented by a group of 31 people without a category and 29 athletes with qualifications. The functional parameters of the cardiovascular system included analysis of heart rate, systolic blood pressure and diastolic blood pressure, which were compared with control values established for the athletes. The immunological indexes were determined in blood serum of the athletes and in subjects from control group by means of the solid-phase ELISA method using conjugated antigens of angiotensin, bradykinin, histamine, dopamine, serotonin for absorption on the plates. To compare the indexes, a control group of 30 people without cardiovascular symptoms was examined. Diastolic blood pressure for athletes of cyclic sports and combat sports was below the control values, and, for athletes of speed-strength sports, it was higher than in control group. A decrease in diastolic blood pressure for the above subgroup of athletes may indicate characteristic adaptive physiological changes in myocardium. The levels of immune-related indexes for serotonin, dopamine for the athletes of all groups corresponded to the control values. The only exception concerned qualified athletes of cyclic sports, which significantly differed from the normal values, compared to athletes without a sports category. Significantly high immunological parameters for histamine and angiotensin did not depend on the athlete’s qualification. In athletes involved in speed-strength sports, the levels of indexes for bradykinin did not differ from the normal values, and, for athletes in cyclic sports and combat sports, they significantly exceeded the norm.

We have shown that different levels and types of sports activities affect the performance of cardiovascular system in their own way. The changes in immunological parameters reflect regulatory state of cardiovascular system. Accordingly, their simultaneous increase against the control may indicate participation in the development of cardiovascular diseases. The differences in their levels for athletes involved in combat sports, cyclic and speed-strength sporting activities show deeper changes in regulatory systems associated with duration and level of physical activities.

Most techniques for evaluation of T-cell immunity are laborious and unsuitable for routine laboratory diagnostics, thus encouraging researchers to look for accessible and reproducible tests. The purpose of our study is to compare three methods aimed for evaluation of cellular immune response levels to the SARS-CoV-2 viral antigens in patients who have been ill and vaccinated against a new coronavirus infection. We have examined 26 persons who experienced mild or moderate COVID-19 (group 1); 19 people vaccinated twice with Sputnik V, who did not have clinical COVID-19 (group 2); 21 subjects who had COVID-19 and were twice vaccinated with Sputnik V (group 3), and 14 persons who had COVID-19 twice (group 4). Peripheral blood mononuclear cells were isolated by gradient centrifugation. The first tested technique was performed as follows: the mononuclear cells were incubated with the S-protein of the SARS-CoV-2 virus, and stained with fluorescently labeled antibodies. The percentage of CD8^highCD107a was counted by means of BD FACS Canto II flow cytometer. When assessed by the ELISpot method with “Human IFN-γ ELISpot” kit, IFNγ production was stimulated by SARS-CoV-2 S-protein, or a mixture of SARS-CoV-2 protein peptides in the “Corona-T-test” kit. There were no significant differences in the levels of CD107a expression on CD8^high cells between the groups 1, 2, 3, and 4, as well as in amounts of IFNγ producers against SARS-CoV-2 S-protein when using “Human IFN-γ ELISpot” kit. Production of IFN was  significantly  lower  in  group  3  (hybrid  immunity), i.e., 317.29±19.04 pg/ml compared to groups 1 and 2 (post-infection and post-vaccination immunity), i.e., 454.95±20.32 and 470.77±26.24 pg /ml, respectively. The relative level of IFNγ -producing cells in group 2 was higher (22.34±3.77) versus 16.83±2.35 in group 1, and 15.46±1.83 in group 3, whereas the relative levels of IFNγ did not differ in these groups. Stimulation with full-length S-protein showed a significant reduction in the number of spots in group 4 (breakthrough immunity), i.e., 30.59±2.29 vs 58.97±4.47 in group 3. Stimulation with a mixture of SARS-CoV-2 peptides in group 4 vs group 3 revealed a significantly increased number of IFNγ -producing cells (86.72±7.20 versus 69.38±5.53) and higher IFNγ production (991.25±65.18 pg/ml versus 760.76±50.70 pg/ml). Appropriate relative values were as follows: 10.30±2.77 versus 8.61±2.66, and 68.10±9.41 versus 48.35±8.15, respectively. The results of three methods for evaluation of cellular immune response correlate positively with each other, but at different significance levels.

The use of dried blood spots (DBS) obtained from the heels of infants has many advantages over the collection of whole blood samples. DNA extracted from DBS can be used to detect genetic diseases by PCR, which has contributed to the development of population-based newborn screening worldwide. Since January 2023, the list of identified diseases includes a group of primary immunodeficiencies (PIDs), associated with the absence or decrease in the levels of T and/or B lymphocytes, determined as part of screening by the levels of TREC and KREC molecules in peripheral blood, respectively. Quantitative analysis requires special attention to biological material. The aim is to evaluate the impact of the preanalytical step on the quantitative analysis of TREC/KREC levels in peripheral blood.

The material included 5219 DBS obtained from infants on days 3-4 of life, as well as DBS prepared from the whole blood of 100 apparently healthy individuals aged 18 to 29 years. A comparative analysis of the TREC/KREC molecules number in correctly and incorrectly collected DBS from newborns and adults, as well as depending on the volume of applied blood, was carried out by RT-PCR using test systems to assess the levels of TREC and KREC in peripheral blood. DBS quality was assessed visually.

In the first months of the project, a significant number of incorrectly taken samples were identified – over a third of all DNA maps received for each corresponding month. As a result of additional training of medical staff, the amount of incorrectly collected material decreased to a level not exceeding 1% of all monthly samples collected. When using DNA extracted from DBS with application errors, the majority of samples (64% for newborns, 78% for adults) failed to obtain a result. In the remaining cases, the results obtained were significantly lower than the normal levels of TREC/KREC determined in the same samples with correct DBS collection (all p < 0.0001, 95% CI). The volume of blood used when correctly applied to Guthrie cards did not affect the results obtained, TREC and KREC levels were comparable; when comparing the medians calculated for each group of samples, no significant differences were identified (p > 0.05).

When quantitatively analyzing TREC/KREC levels in peripheral blood, correctly taken material is fundamental importance to obtain reliable indicators, primarily to exclude false-positive results. To minimize errors in the preanalytical stage, additional training of medical personnel is necessary to control and/or correct errors.

Neonatal screening is a mandatory newborn screening procedure that detects the presence of genetic diseases. Dry blood stains are used for mass screening of children. This technology is the most affordable and convenient for the transportation and storage of biological material. DNA extraction is one   of the important steps in molecular diagnostics, the accuracy of which is particularly important in genetic analysis. Different DNA extraction kits offer different protocols and reagents, which may vary in efficiency and quality of extraction.

The aim of our work was to perform a comparative analysis of reagent kits for DNA extraction from dried blood spots.

The materials were dried blood drop samples on Guthrie cards obtained from healthy preterm infants on day 3-4 of life as part of a newborn screening program.

The study methods included spectrophotometric analysis to determine the concentration and purity of DNA, the simplicity of protocols, the duration of isolation, and the possibility of automating the process were also evaluated. The efficiency of DNA isolation using different reagent kits was additionally monitored by real-time PCR results using a test system to assess the level of TREC and KREC in peripheral blood, since quantitative analysis requires more attention to the material under study.

In assessing the purity of the nucleic acid extracted using four kits analyzed, successful deproteinization of DNA samples and relative purity could be observed. The average DNA purity for the “Extra-DNA-Bio” set was 2.2±0.23, for “EXTRA-prep PS” – 1.89±0.23, for “MagnoPrime UNI” with manual and automatic isolation – 2.31±0.21 and 2.85±0.09, respectively. The average DNA concentration for the Extra-DNA-Bio kit was 15.28 μg/mL, for the EXTRA-Prep PS kit it was 16.26 μg/mL, and for the MagnoPrime UNI for manual and automated isolation it was 62.5 μg/mL and 102.28 μg/mL, respectively. According to the applied Kraskell-Wallis criterion and Dunn’s test, significant differences in both TREC and KREC parameters are present between the group of DNA samples extracted using the “MagnoPrime UNI” reagent kit for manual extraction and the groups of samples extracted by other methods (“MagnoPrime UNI” for automatic extraction) or “Extra-DNA-Bio” and “EXTRA-Prep PS” kits.

In the course of the present study, four comparable reagent sets demonstrated a high level of convergence of the obtained data, satisfying all the necessary parameters for further molecular genetic analysis, can be used for neonatal screening and in other areas of research requiring DNA extraction from a dried blood spots.