Immune checkpoints represent the system of inhibitory mechanisms regulating the activation of the immune response, preventing the autoimmune processes and modulating the immune response by decreasing the immune cell-mediated damage of tissues and organs. Tumor cells may utilize these checkpoints to prevent the activation of tumor-specific lymphocytes, thereby acquiring resistance against the immune response. The blockade of inhibitory signal that is transduced in immune checkpoints leading to the reactivation of antitumor immune response is a promising method of tumor immunotherapy. Since the majority of immune checkpoints are based on the ligand-receptor interactions, one of contemporary modalities of anti-tumor therapy is based on the development of ligandor receptor-blocking therapeutic monoclonal antibodies, as well as soluble recombinant receptors capable of competing for a ligand and thereby modulating the signal transduction. In the past few years, this field of tumor immunotherapy experienced an impressive success; however, the potential tradeoff for altering of the natural suppressive mechanisms is the development of the autoimmune reactions.

Allergen-specific immunotherapy (ASIT) is the most effective method of allergy treatment which consists of exposure to small doses of antigen responsible for development of allergic condition in the particular patient. Therefore, one may achieve desensitization to this antigen. The history of ASIT application lasts for more than 100 years, and, over this time, huge clinical evidence for the usage of the method has been accumulated. Use of ASIT causes reduction of allergy symptoms and treatment needs and, moreover, it has the potential for long-term clinical benefit, by preventing the development of allergy and its symptoms. The treatment affects basic immunological mechanisms responsible for the development of clinical symptoms. ASIT is an antiinflammatory, pathogenetic and prophylactic treatment of allergic airway disease. The review considers the results of major clinical trials of the ASIT applications for treatment of allergic diseases of the respiratory system (allergic rhinitis and bronchial asthma). Various schemes of ASIT are discussed including its different variants (injectable and sublingual ASIT), the issues of preparation choice for ASIT from those currently available on the pharmaceutical market, patient selection criteria, and the issues of modern molecular allergodiagnostic (allergic sensitization mapping of the patient at molecular level), in order to optimize them. Immunological mechanisms of ASIT are also considered, since appropriate views are rather contraversial. The ASIT effect is mediated through the following basic immunological mechanisms: the suppressed increase of the eosinophil concentrations, reduced duration of the delayed hypersensitivity phase, as well as initiation and maintenance of the Th2-to-Th1-like immune response transition. Regulatory T-cells play a major role in implementation of the immunological mechanism in ASIT, they have a significant impact on the Th2 response suppression. Such suppression may proceed via increased production of specific IgG antibodies, primarily of IgG4 subtype. The shift of balance between IgE and IgG4 towards IgG4 production is now considered a fundamental condition for successful ASIT. It has been proven that the allergen-specific IgG4 antibodies against IgE persist after discontinuation of the treatment and can provide long-term clinical tolerance. Modern prospectives for development of new forms and species of preparations for ASIT are also reviewed. Two groups of drugs for ASIT have been adopted for clinical practice until now, i.e., allergens and allergoids (allergens chemically modified by treatment with formaldehyde, in order to enhance their immunogenicity and to reduce the incidence of adverse allergic reactions associated with their application). Immunological mechanisms of SLIT (sublingual immunotherapy) are subjected to special consideration. So far, SLIT is the safest and most promising option of ASIT today, its usage is most expedient in pediatric practice.

Latent rhinosinusitis proceeds without facial pain symptoms. Immune deficiency plays a leading role in pathogenesis of the disease latency. Substance P seems to be a universal mediator of painful irritation and inflammation. The objective of our study was to determine effectiveness of therapies in patients with latent rhinosinusitis, in terms of substance P levels.

We treated 148 patients with rhinosinusitis, being free of local pains. All the patients underwent clinical and laboratory examination, including immune profile assessment, measurements of serum cytokines IL-1β, IL-4, IL-6, IL-8, IL-10, TNFα, IFNγ, and substance P. To correct a secondary immunodeficiency, the standard treatment of rhinosinusitis in a subgroup of the patients was accomplished by immunomodulatory drugs from the first day of therapy. The latter drugs were avoided for the rest of study group. Efficacy of treatment was evaluated by clinical signs and laboratory parameters on day 7 of the medication.

 Pre-treatment levels of substance P were determined in all the patients with latent clinical course and lack of pain symptoms. Low substance P levels (< 100 pg /ml) were considered as indications for immunomodulatory therapy, due to immune deficiency confirmed by the cytokine imbalance. Choice of a specific drug was dependent on immunopathogenesis, i.e., for catarrhal rhinosinusitis and deficiency of cellular immunity, we administered IFN-ES-lipint; in cases of purulent rhinosinusitis, Likopid was applied. The patients treated with immunomodulatory drugs showed improvement of immune indexes by the 7th day of treatment, along with return of substance P levels to control values typical to healthy persons. Among patients with low substance P levels and immune deficiency (without immunomodulatory treatment), the immune parameters and substance P levels did not exhibit any sufficient changes over time.

Low contents of substance P (SP ≤ 100 pg /ml) in blood serum in pain-free patients with latent rhinosinusitis are indicative of immune deficiency and may serve as an indication for immune modulation therapy. Individual selection of the pathogenetically proven schedule therapy, when treating patients with latent (painless) rhinosinusitis may result into effective prevention of severe inflammation, normalization of immune response, prevention of a protracted disease course, and appropriate complications, as well as shortening of treatment terms to 8-10 days

The aim of this study was to evaluate subpopulational profiles of Tand B-lymphocytes in the patients with chronic obstructive pulmonary disease (COPD) under the influence of a multivalent vaccine against Pseudomonas pathogens, in order to justify its usage as a potential immune stimulator of microbial origin. Sixty patients (39 males and 21 females aged 40 – 58 years old) with verified diagnosis of COPD II-III degree were included into the study group. All the patients received a standard COPD therapy. In accordance with planned treatment, this cohort was divided into two groups using a simple randomization. Group 1 included thirty COPD patients (20 men and 10 women) who received standard treatment. Group 2 consisted of thirty COPD patients (19 men and 11 women) who received treatment according to conventional schedule, but further, they were additionally treated with «Pseudovac» vaccine used as a infection-preventing measure. The «Pseudovac» vaccine was injected, according to appropriate instructions for the drug use. Immunological evaluation was performed three times: at admission, as well as 10 and 20 days of treatment. As a control group, 35 healthy individuals (19 men and 16 women) of a similar age range have been examined. Assessment of Tand Bcellspecific immune phenotypes was determined by means of flow cytometry using direct immunofluorescence of leukocytes whole peripheral blood using specific monoclonal antibodies. We have found that Tand Blymphocyte profiles in the COPD patients are characterized by decreased numbers of T-cells (due to drop in cytotoxic T-cells), and increased B-lymphocyte contents. Relative amounts of NKTand γδT-lymphocytes showed an increase in cases of developing bacterial infection. The T-lymphocyte subpopulations in patients with COPD are characterized by increase in relative numbers of regulatory T-cells, Th2-lymphocytes, and T-cells expressing activation markers. B-lymphocyte population in COPD patients was characterized by increased levels of B2-, B1and memory B-cells, along with reduced levels of CD23-receptor-positive B-lymphocytes. During a standard treatment of infectious and inflammatory disease in COPD patients, there is a tendency to normalization of Tand B-lymphocytes subpopulations. However, T lymphocyte number by the 20th day of treatment proved to be reduced (due to decrease in cytotoxic cells), whereas B-lymphocyte content was increased. Furthermore, the levels of NKTand T-regulatory cells remain high at the standard COPD treatment. The «Pseudovac» vaccine exerts a pronounced immunoactive effect upon adaptive immunity, by means of restoring and/or normalizing Tand B-lymphocyte subpopulations in COPD patients by the 20th day of vaccine treatment.

Modulatory effects of three probiotic bacterial strains (Lactobacillus rhamnosus K32 (L), Bifidobacterium longum GT15 (B, Enterococcus faecium L-3 (E) on expression level and contents of key cytokines were studied using PCR techniques with reverse transcription, and enzyme-linked immunosorbent assay. Both cell cultures and an experimental model of intestinal dysbiosis were used in this study.

The genes encoding bacteriocins, surface membrane component, pili and exopolysaccharides involved in host immune system modulation were previously identified in the B and Ebacterial strains.

Investigation of probiotic strains and effects of their supernatants expression of cytokines in cell cultures of promonocyte origin (HTP-1) showed increased expression of TNFα, due to E and L supernatants. Moreover, the Bl culture induced IL-8 and IL-10 expression.

In a model of Wistar rats with ampicillinand metronidazole-induced intestinal dysbiosis corrected with probiotics we have shown that the dysbiosis was accompanied by sufficient alterations in microbiota composition (Klebsiella spp. overgrowth and low contents of Faecalobacterium prausnitzii) that were observed only in the animals untreated with probiotics (control), or after administration of L.

In contrast to these results, the animals treated with E and B, the following changes were revealed: 1) low expression of proinflammatory cytokines IL-8, TNFα, MCP-1 inmesenteric lymph nodes and appropriate changes of their serum contents, 2) increased serum content of the anti-inflammatory TGFβ cytokine. Hence, the present study, having used two complementary models, has detected some individual features of immune modulation produced by the probiotictic strains of L. rhamnosus K32, B. longum GT15 и E. faecium L-3 which exert differential effects upon the intestinal microbiota. 

Stress-induced immune disregulation is a risk factor of autoimmune and inflammatory diseases, but, so far, the mechanisms for this effect are not fully known. Expression levels of specific mRNAs were assessed in gut-associated lymphoid tissue (GALT) from Wistar rats subjected to chronic social stress (CSS). Gene expression was evaluated for NR3C1, Adrβ2, as well as IL-1β, IL-17α pro-inflammatory cytokines, and Nlrp, an inflammasome gene. Under the CSS conditions, we have shown altered distribution of RORγt +, FoxP3+, LMP2+, XBP1+ lymphocytes in GALT.

The experiments were carried out with female Wistar rats aged 5–6 months. Specific mRNA expression for the target genes was determined by means of real-time PCR performed in a CFX96™ thermocycler («BioRadLaboratories, Inc»,USA). Relative levels of a target gene expression were quantified by the ΔΔCt method, being compared with rat GAPDH reference gene expression. Statistical analysis was performed with available «BioRad СFX Manager 3.1» software. Specific monoclonal rat antibodes were used for detection of immunopositive lymphocytes by means of indirect immunofluorescence technique.

CSS development leads to decreased levels of mRNA expression for Nr3c1 and Adrβ2-genes in the GALT cells, being accompanied with unidirectional changes, i.e., increased transcription of pro-inflammatory cytokine mRNAs (IL-1β, IL-17α) and Nlrp3-inflammasome genes. These changes are accompanied by decreased FoxP3+/RORγt + cell ratio and predominant Th17 differentiation accompanied by suppressor failure. In addition, CSS development was characterized by unidirectional tendency for increasing total number of LMP2+ lymphocytes and reduced ХВР1+ cell population density in lymphoid structures of rat ileum.

The events observed in GALT cell populations under CSS conditions are opposing classical paradigm of the stress response. The CSS-associated effects do not promote immunosuppression, however, are able to cause activation of immune system and inflammatory process. 

The article presents results of studies concerning subрopulations of peripheral blood lymphocyte in the patients at early stages of age-related macular degeneration (AMD), being compared with healthy elderly persons (risk group for AMD), and young adults without ophthalmological problems. We have revealed an increase in absolute counts and percentages of the cytotoxic cells (СD3+СD8+), and double-positive T cell subpopulations (СD3+CD4+СD8+), like as of B-lymphocyte contents, higher frequency of their increased content in peripheral blood of the patients with early and intermediate stages of AMD and healthy older people, as compared with young controls, thus allowing to suggest a potential role of these shifts in lymphocyte subpopulations for the AMD pathogenesis.

Primary immunodeficiencies (PID) such as severe combined immunodeficiency (SCID) and X-linked agammaglobulinemia are characterized by the lack of functional Tand B-cells, respectively. Without early diagnosis and prompt treatment children with PID suffer from severe infectious diseases, leading to their death or disability. Our purpose was developing of simple, inexpensive, high throughput technique based on the quantitative determination of TREC and KREC molecules by real-time PCR, and its validation in a group of children with a verified diagnosis of SCID and X-linked agammaglobulinemia.

In this study, we developed and validated multiplex real-time PCR for the TREC’s and KREC’s quantitative analysis. We have shown that linear range of Ct changes depending on the concentrations of targets with a correlation coefficient R2 not worse than 0.98 was observed at concentrations from 109 to 5 × 104 copies per ml. The lowest amount of targets reliably detected in a reaction volume was 10 TREC’s copies, 5 KREC ‘s copies and 5 copies of internal control (IL17RA). We determined the age-depended reference values of TRECs and KRECs in whole blood in 29 boys and 27 girls with normal immunological parameters. The normal cut-offs for TRECs and KRECs were defined in dry blood spots depending on the method of extraction.

The proposed method showed 100% diagnostic sensitivity and specificity in the studied group. The method can be proposed as a screening tool for the diagnosis of SCID and X-linked agammaglobulinemia both in whole blood and in the dry blood spots. The further investigation is required with larger number of samples. 

Complement system is an important component of innate immunity, providing primary protection against pathogens invading the body. In addition, it was shown that the complement system is associated with many diseases, not only autoimmune and infectious, but also mental disorders. In this regard, it is necessary to develop affordable and fast method of measuring activity of the complement system in real-time mode. We present a new semi-automated method for assessment of serum complement activity. The assay is based on cytolytic action of complement system upon the ciliate organism Tetrahymena pyriformis. This method consists in repeated counting of live Tetrahymena motile cells by means of specially developed Biolat device, which consists of two video cameras, light sources, and movable round plate. The plate has two rows of holes. The device also includes microprocessor control unit based on AutoCiliata software, intended for control of operation module and counting the surviving cell. The calculations are based on fixation of two sequential video-frames, with subsequent software image processing. Cell death events were observed upon incubation in triethanolamine (TEA) buffer containing 5% of blood serum. We have also compared complement activity in different buffers, i.e., standard medium for culturing of ciliates, Veronal-Medinalum buffer, and the TEA buffer. TEA buffer was found superior to the Veronal buffer when applied in the test system. The time of cell death in the TEA-buffered medium containing 5% serum was < 15 minutes for all the sera studied. The parameters denoting serum complement activity were as follows: a half-life time for the moving cells (TLD50), and a similar value for 100% cell inactivation (1/TLD50, functional activity of the complement system, ACS). The sensitivity of this assay was calculated from dependencies between TLD50 and ACS, and actual serum concentrations. We have suggested an opportunity for evaluation of an integral complement activity, and interrelations between the intensity of synthesis and consumption of its major effector proteins. In the course of this study, we have tested different concentrations of Ca++ and Mg++ ions in the incubation buffer, with optimal physiological concentrations of2.5 mMand1.5 mM, respectively. We have also estimated statistical precision characteristics for pre-analytical and analytical steps of the method. The average coefficients of variation (CV) were 3.9% and 2.7%, respectively, thus satisfying the reliability criteria in research. A short performance time of the study suggests its potential application in clinical practice, including online examination regimens. A method for semi-automatic measurement of serum complement activity could be applicable in daily clinical practice, including the online performance.