Thymic mast cells: at three-way crossroads
The review examines the role of thymic mast cells in the interaction of three integrative systems - nervous, endocrine and immune. Mast cell structure, the mechanism of connections with nerve terminals, the spectrum of mediators and hormones that provide intercellular communication are analyzed. It has been shown that mast cells are both receptor and effector cells that enhance the mechanisms of nervous regulation. The review also provides arguments in favor of mast cell contribution to the thymus endocrine function. The role of thymic mast cells in the cooperation of three integrative systems is considered in the context of regulation of T-cell maturation and differentiation.
Study of the immune response and metabolism state using experimental models of hyperlipidemia and metabolic syndrome
Introduction. The immune system plays an important role in the pathogenesis of hyperlipidemia (HL) and metabolic syndrome (MS). Metabolic dysfunctions are accompanied by disorders of cooperation, intercellular interaction of various parts of the immune system. The key pathogenetic factors of GL and MS are changes in the secretion of adipocyte mediators (adiponectin, leptin), pro-inflammatory and anti-inflammatory cytokines (CT). The data of experimental and clinical studies indicate the relationship and interdependence of cellular immune responses, mediators of intercellular interactions and metabolic disorders of the macroorganism.
The aim of this study was to estimate the population composition of spleen lymphoid cells, the production of cytokines and the functional activity of lymphocytes in 2 experimental models of MS and HL.
Material and methods. Experiments were performed on C57Bl/6 mice. The studies were carried out on two experimental models of MS and GL, based on long-term feeding of animals with a 20 % aqueous solution of fructose with the addition of cholesterol to the food and intraperitoneal administration of Poloxamer 407 to mice, respectively. The subpopulation composition of mice spleen cells was assessed by flow cytometry. Determination of leptin, adiponectin, insulin, cytokines interleukin(IL)-15, IL-22 in the blood serum of mice was performed by ELISA using commercial kits. The proliferation of lymphocytes was evaluated in the blast-transformation assay (RBTL).
Results. The results of the experiments carried out in the MS and HL models indicate a change in the population composition of the spleen (a decrease in CD4+- and CD8+-T cells numbers, activation of CD4+CD25+Foxp3+-Treg cells), accompanied by a decrease in the functional activity of immunocompetent T cells and an increase in the proliferative activity of B-lymphocytes. Also a violation of the production of IL-15 and IL-22, involved in lipid metabolism and dysfunctional changes in the mediators of lipid and carbohydrate metabolism (adiponectin, leptin, insulin) was observed, which in turn may serve as a prerequisite for the development of chronic inflammation, considered as the most important pathogenetic sign of MS.
Conclusion. Our experimental data on changes in the immune response and the state of metabolism confirm the feasibility of searching and screening for immunocorrection drugs intended for the prevention and treatment of hyperlipidemia and the MS symptom complex, based on a targeted effect on the leading steps in the pathogenesis of metabolic disease.
Dynamics of T-lymphocyte subpopulations’ recovery after influence of various damaging agents
Introduction. Various damaging agents have different immunosuppressive effect on the immune system and cause pathogenetically varied reparative changes in it. Studying the mechanisms of T-cell recovery is important both for understanding the pathogenesis of reparative processes and for finding adequate methods for their correction.
Aim - to study the role of thymus and peripheral lymphoid organs (for example, the spleen) in the T-lymphocyte restoration after equal damage by cytostatic cyclophosphamide or sublethal irradiation.
Material and methods. C57BL/6 mice were divided into 3 groups. Group 1 (20 mice) received with cyclophosphamide once, group 2 (20 mice) was irradiated with X-rays (4 Gy) once, and group 3 (8 mice) was used as control. Mice were euthanized on days 10, 20, 30 and 60. The cellularity and subpopulations of T-cells in thymus and spleen were determined by flow cytometry.
Results. We have shown that radiation damages actively dividing thymocytes more deeply than cyclophosphamide. Some block is formed in the thymus at the stage of DN4 thymocytes, and complete recovery of cytotoxic T-cells in the spleen is not observed even by two months after sublethal irradiation. Radiation is more damaging to the CTL subpopulation, and CTLs are not completely restored either in the thymus or in the periphery by day 60. Actively dividing thymocytes are more susceptible to the action of damaging factors than mature splenocytes. Homeostatic proliferation with the conversion of the phenotype of naive T cells into central memory T-cells makes a significant contribution to the regenerative processes.
Conclusion. T-cells are more sensitive to ionizing radiation than to the action of cytostatics. And irradiation significantly damages not only the lymphoid elements, but also the thymic stroma. Both types of exposure lead to a decrease in the number of naive T-cells and the accumulation of surrogate memory T-cells, which should result in a narrowing of the T-cell receptor repertoire, the development of immunodeficiency and immunopathology associated with the T-cell response. We should understand it when prescribing the optimal corrective treatment after radiation and chemotherapy.
Local cytokine status in patients with sialadenosis of parotid glands with hypothyroidism
Introduction. The salivary glands’ dystrophic and inflammatory diseases occupy a significant proportion of the prevalence of dental pathology. It is known that the pathogenesis of sialadenosis is associated with thyroid gland dysfunction. Local treatment of sialadenosis and correction of somatic disease is not enough, as it’s not possible to achieve in these patients stable remission. It has been experimentally proved that already in subclinical forms of hypothyroidism there are morphological changes in the parenchyma of parotid gland with lymphoid infiltration. The study of local cytokine levels can largely determine outcome of the disease.
Aim - to study pro- and anti-inflammatory cytokine levels in parotid secretion of patients with sialadenosis and hypothyroidism.
Material and methods. The study consisted of 52 patients with sialadenosis of parotid glands. All the patients were divided into 2 groups: 1st - 30 patients in remission stage, 2nd -22 patients in the acute stage. The comparison group were included 20 healthy volunteers. The content of IL-1β, IL-6, TNFα and IL-10 was determined in the parotid secretion by enzyme immunoassay (ELISA) using Vector-Best reagents (Russia) and BMG Labtech (ClarioStar, Germany).
Results. There was a significant difference in the levels of IL-1β, IL-6, TNF-α between the groups of sialadenosis patients and the group of comparison with the maximum cytokines’ level under study in sialodenosis in the acute stage. The level of IL-10 was significantly lower in sialodenosis in the acute stage compared to the other groups. TNFα was detected in parotid secretion only in 25 % of subjects in the group of comparison.
Conclusion. The study of proinflammatory and anti-inflammatory cytokines in the salivary gland secretion, along with the clinical picture, can serve as an additional diagnostic tool to determine the severity of the inflammatory process in these patients, which allows to consider the use of this method in practical dentistry for dynamic monitoring of patients and further application of treatment efficacy.
Analysis of subpopulations of bone marrow lymphocytes in patients with acute myeloid leukemia: immunological effects of chemotherapy
Introduction. Acute myeloid leukemia is one of the most common types of blood cancers with low five-year survival rate and poor prognosis. The gold standard of treatment for this disease is a chemotherapy scheme called «7+3» which stands for a combination of cytarabine and an anthracycline antibiotic, usually daunorubicin. Lymphocytes play a pivotal role in cancer immunosurveillance. In this study, we conducted a detailed analysis of the effects of the «7+3» chemotherapy regimen on bone marrow lymphocytes and examined several parameters of lymphocytes from patients with acute myeloid leukemia.
Aim of the study is to investigate the percentage and phenotypic characteristics of bone marrow lymphocytes in AML patients at different stages of «7+3» chemotherapy course.
Material and methods. Bone marrow aspirate samples were obtained from 61 AML patients. Among them, 16 patients were treated according to the classical «7+3» scheme and had at least two samples before and after therapy. Erythrocyte lysis buffer was used to isolate leukocytes from bone marrow aspirates, then cells were stained with a panel of antibodies, which included anti-CD3-, CD4-, CD8-, CD16-, CD19-, CD45-, CD56- and CD57-antibodies. FACS analysis was performed on a BD Aria II flow cytometer and data were analyzed using FlowJo software.
Results. The ratio of lymphocyte populations changed during the therapy in the group of 16 AML patients. We found out that B cells were extremely sensitive to «7+3» therapy; they were eliminated by the early post-therapeutic period. NK cells began to recover 2-3 weeks after the end of the course of chemotherapy. Within the total group of 61 AML patients the ratio of CD4+/CD8+ T cells was slightly increased, although this parameter stayed within the normal range. 3 patients whose NK cells did not express CD16 were identified, however, this phenotype did not affect the effectiveness of chemotherapy and patient outcomes.
Conclusion. The present study revealed patterns of changes in the ratio of lymphocyte populations and their surface phenotype under the influence of the «7+3» chemotherapy regimen. In addition, the lack of expression of the surface molecule CD16 on the NK cells of some patients was demonstrated.
Clinical and immunological correlations in COVID-19: experience of immunomodulatory therapy
Introduction. Attempts to influence the interaction between SARS-CoV-2 and immune system provide additional opportunities to prevent both COVID-19 development and progression of its complications.
Aim of the study - to evaluate the reasonability of immunomodulatory therapy in the complex treatment of hospitalized COVID-19 patients.
Material and methods. 43 hospitalized patients with COVID-19 were included in the study: comparison group (n = 21) and the main group (n = 22), in which on the 7-14th day of hospitalization after standard complex treatment was additionally used a protease inhibitor with virocidal properties, aprotinin, aminodihydrophthalazinedione sodium (Galavit®), normal human immunoglobulin (IgG, IgA, IgM), interferon alpha-2b.
The following indicators were assessed at 1-3, 5-7 and 9-15 days from the start of the immunomodulatory therapy: level of interleukin(IL)-6, T cells (T-helpers and cytotoxic T-lymphocytes), immunoregulatory index (СD4+/СD8+ ratio). The indices were measured by EIA, cytofluorimetric and immunochemiluminescent methods.
Results. In the main group pro-inflammatory cytokine IL-6 level significantly decreased against the background of immunomodulatory therapy, IL-6 level changes directly correlated with the use of immunomodulators. In the main group, the level of T-cell subpopulations began to recover starting from the 3rd day of observation, reaching the lower normal limit by the 14th day of treatment with immunomodulators. After 7 days of observation, in 18 (81.8 %) patients with SARS-CoV-2 in the main group CD4+/CD8+ ratio was ≥ 1.4. In the comparison group, 15 (71.4 %) patients with moderate disease course had a CD4+/CD8+ ratio ≤ 1.4 and 2 (9.5 %) patients had CD4+/CD8+ ratio of ≥ 1.4. Viral load decreased significantly faster in the main group (p ≤ 0.005). According to PCR tests, SARS-CoV-2 elimination occurred in the main group at day 11 in all patients, and in the comparison group, the PCR test was negative only in 4 out of 20 patients (25 %) on the 21st day of the disease. АП patients in the main group showed positive clinical dynamics. In the comparison group, despite the standard treatment carried out, no significant positive dynamics was achieved; noted the addition of a secondary bacterial infection on the 21st day of the disease.
Conclusion. Thus, immunomodulatory therapy may improve the effectiveness of treatment of patients with COVID-19.
Dynamics of immunological and microbiological indicators of oral fluid in caries therapy
Introduction. The mucous membrane of the oral cavity is part of a single system of mucosal immunity and implements functions at the level of local and systemic immunity. Microflora together with innate immunity receptors (TLR, NLR, etc.) regulates the colonization resistance of mucous membranes, determines the severity of the infectious process and maintains the normocenosis of the oral cavity. Violations of the dynamic balance of colonization resistance can lead not only to dental caries, but also to other pathologies of both the oral cavity and the whole organism.
The aim of this study was to estimate the levels of α-defensins and secretory immunoglobulin A (sIgA), as well as representatives of the microbial community of the oral cavity of young people aged 19-22 years with caries of the occlusal and contact surfaces of posterior teeth against the background of prophylactic use of Licopid® 1 mg (glucosaminyl muramyl dipeptide, GMDP).
Material and methods. In 43 clinically healthy patients 19-22 years old with caries of the occlusal and contact surfaces of the lateral teeth, oral fluid was taken before taking the drug and 14 days after. Patients of the main group of 21 people from 1 to 10 days of observation received drug Licopid® (1 mg) (GMDP, AO Peptek, Moscow, Russia) sublingually, 1 tablet once a day. Participants of the comparison group (22 patients) did not received Licopid®. Oral fluid microbiota was assessed by RT-PCR using Dentoflor reagents («DNA-technology», Moscow). Primers V6 16S rDNA were used for NGS DNA sequencing on an MiSeq instrument (Illumina, USA). Microorganisms were identified using the Blast software. The analysis of α-defensins HNP1-3 and the content of sIgA in sublingual saliva were studied by enzyme-linked immunosorbent assay using Hycult Biotech (USA) and Vector-Best (Novosibirsk, Russia) reagents. For statistical processing of the data the Mann-Whitney test was used and the Biostat software.
Results. The use of the drug Licopid® 1 mg helps to restore the levels of α-defensins HNP1-3 and sIgA to the level of those in individuals with intact teeth, leads to a significant decrease in the representation of Porphyromonas gingivalis and Clostridium difficile and increases the diversity of oral microflora.
Conclusion. The use of Licopid® 1 mg after the caries treatment is accompanied by the correction of immunological parameters in the oral fluid.
The use of the ELISPOT technological platform in the diagnosis of tuberculosis infection in patients with HIV infection
Introduction. The problem of the steady growth of HIV-associated tuberculosis has become more and more urgent in recent years in Russia. Diagnosis of tuberculosis is especially difficult at the stage of severe immunodeficiency.
Aim of the study - to assess the possibility of using the T-SPOT.TB method in the diagnosis of tuberculosis infection in patients with HIV infection.
Material and methods. In the period for 2019-2020, an analysis of the T-SPOT.TB results was carried out in 396 patients. The inclusion criteria for this study were age 18 and older, diagnosis of HIV infection, dispensary observation at the AIDS center, CT scan of the lungs, and the presence of a blood test for T-SPOT.TB. Based on the results of the T-SPOT.TB test, 2 groups were formed: group 1 - with a positive result - T-SPOT.TB+ (n = 143) and group 2 with a negative result - T-SPOT.TB- (n = 253).
Results. Tuberculosis was verified by detection of Mycobacterium tuberculosis in 117 patients with HIV-infection, all CT changes characteristic of tuberculosis, a positive T-SPOT. TB result was in 88 % (n = 103) cases. The mean CD4+ T cells level in this group was 230.05 ± 15.3 cell/mcl, 95 % CI [190.91- 256.2]. The diagnosis «latent tuberculosis infection» was made in 40 patients with HIV infection with a positive T-SPOT.TB result in the absence of changes on CT.
Conclusion. T-SPOT.TB can be used in a diagnostic complex for monitoring patients with HIV infections as a screening method that allows not only to verify the diagnosis in the future, but also to reveal latent tuberculosis.
Analysis of the mechanisms of development of the immune response in hepatitis B virus infection and ways to improve the effectiveness of vaccination
Currently, a number of recombinant hepatitis B vaccines with a proven immunogenicity and safety profile are used in medical practice. Vaccination against this infection within the framework of national immunization programs in different countries of the world, including in Russia, helped to reduce the incidence of hepatitis B. However, despite the availability of effective drugs, there are groups of people who do not respond well to vaccination. Since the severity of the immune response to the hepatitis B virus is determined by many factors, an active search for approaches to improve the effectiveness of immunoprophylaxis in this infection continues. The article discusses the main ways of developing immune responses in hepatitis B virus infection, provides information on ways to improve the effectiveness of vaccination against this pathogen, one of which is the use of adjuvants. A brief description of the registered hepatitis B vaccines containing various adjuvants is presented. The main mechanisms of their action are considered; the results of a number of preclinical and clinical studies of modern hepatitis B vaccines using new adjuvants, which may be promising in terms of increasing the immunoge-nicity of vaccines, are summarized. Understanding the features of the formation of immunity against the hepatitis B virus in combination with the study of the mechanisms of action of new adjuvants will contribute to the creation of modern drugs that provide a balanced stimulation of the humoral and cellular immune response against the hepatitis B virus.
Development and function of regulatory B cells and its role in pregnancy support
Traditionally, B cell function is predominantly associated with humoral immunity and positive regulation of the immune response through the production of specific antibodies and optimal activation of CD4+-T cells. However, a unique subclass of B cells has recently been identified, later described as regulatory B cells (B-regulatory, Breg). These cells are capable of suppressing the functions of other immune cells, including dendritic cells (DC) and T-lymphocytes, through the production of anti-inflammatory cytokines, predominantly interleukin(IL)-10, which effectively inhibits the proliferation and production of pro-inflammatory cytokines by T cells and induces the differentiation of regulatory T cells (T-regulatory, Treg cells). Immunosuppressive Bregs that produce IL-10 are involved in maintaining homeostasis in the immune system and self-antigen tolerance. The data obtained confirming the protective role of Bregs in the completion of inflammation, as well as their pathogenic role through the production of autoantibodies. The Breg population includes a unique functionally defined subclass that can currently only be detected by its ability to produce and secrete IL-10 following appropriate ex vivo stimulation. Evidence is now accumulating to support an important role for Bregs in autoimmune and infectious diseases, as well as in the formation of graft tolerance and tumor development. Breg are also involved in the formation of selective immune tolerance during pregnancy, which is necessary to ensure the immuno-privileged microenvironment of the semi-allogeneic fetus. It has been shown that a violation of the number and functional activity of Breg can lead to the development of various immunologically mediated pathologies of pregnancy, such as recurrent implantation failure, recurrent pregnancy loss and premature birth. This review presents the results of studies on the biology and functional role of Breg in the regulation of reproductive processes.
Antimicrobial proteins and peptides of neutrophilic granulocytes as modulators of complement system
Neutrophilic granulocytes (neutrophils) and complement system are important factors of the innate immunity of vertebrates. Due to a long coevolution different interactions between them have been established. The data about action of antimicrobial proteins and peptides of neutrophils on complement have been collected to date. In this paper, the biology of neutrophils and complement is briefly considered and research reports are reviewed to systematize data about action of myeloperoxidase, reactive oxygen species, lysozyme, antimicrobial peptides, lactoferrin, neutrophilic extracellular traps, neutrophil fragments and total matter on human complement. Physiological and pathophysiological role of this modulation is emphasized.