The present review contains data from literature concerning the in vivo structure of synovial membranes in healthy people and patients with rheumatoid arthritis (RA). The properties of in vitro cultured fibroblast-like synovial cells (FLS) from RA patients are considered, including FLS morphology, phenotype and function. A standard protocol of in vitro FLS culturing is described. Notably, the FLS are characterized by autonomic functioning, ability for invasive growth/migration, e.g., into non-affected joints. These FLS properties may a reason of multiple joint involvement typical to RA. Special attention is drawn to characterization of stable phenotypic profile of FLS which results from certain epigenetic disturbances, i.e., changes of the DNA methylation, histone acetylation, and micro-RNA effects.

The FLS from RA patients are characterized with stable and extensive hypomethylation of genes which occurs in vivo and persists after repeated culture passages. Some promoters of genes involved into RA pathogenesis (for example, CXCL12, IL-6) are hypomethylated. By contrary, some other gene promoters (e.g., the death receptor 3 gene) are shown to be hypermethylated. An increased histone acetylation of genes encoding proinflammatory mediators (such as MMP1) may be an important mechanism of persistent inflammation in RA. Changes in histone acetylation in FLS are related to high levels of ubiquitin-like SUMO-1 protein and concurrent decrease in specific protease SENP1activity. A role of histone acetylation in RA pathogenesis is supported by efficacy of a histone deacetylase inhibitor (Trichostatin A) in collagen-induced murine arthritis. Local concentrations of micro RNA-155, micro-RNA-146а, and micro-RNA-203 are permanently increased in FLS cultures, synovial tissues, and PBMC of the RA patients. Expression of micro RNA-124а is decreased in FLS from RA, as compared with OA FLS.

One may conclude that the fibroblast-like synovial cells are key cellular elements involved in the pathogenesis of rheumatoid arthritis, and the studies of their impaired epigenetic regulation are at their initial stage. Enzymes and molecular complexes involved in these processes may represent potential therapeutic targets for the treatment of RA.

Thymectomy is a stage of surgery when treating some congenital heart defects. Thymus gland is the central organ of immune system. This organ is the primary site of T-cell lymphopoiesis and central tolerance to autoantigens during fetal and early postnatal life. If performed neonatally or in infancy, the thymectomy may cause restriction of these immune functions. Suppression of T-cell lymphopoiesis in children with thymectomy can be estimated as a subpopulation of thymic naive T helper cells (CD3+CD4+CD45RA+CD31+). To perform this task, we evaluated subpopulations of thymic naive T helper lymphocytes with CD3+CD4+CD45RA+CD31+ phenotype in the children (n = 40) who underwent thymectomy during surgical treatment of congenital heart diseases in neonates, or in early postnatal life. Their data were compared with children who underwent surgical treatment of congenital heart disease without thymectomy at the same age periods (n = 12), and healthy children (n = 23). We have revealed that thymectomy in frames of surgery of congenital heart disease leads to reduced thymic naive T helper lymphocytes with CD3+CD4+CD45RA+CD31+ phenotype in peripheral blood. Early execution of thymectomy is associated with deficiency of the thymic naive T helper lymphocytes in the peripheral blood, as well as a decrease in T helper cells (CD3+CD4+). The number thymic naive T helper lymphocytes in peripheral blood negatively corrrelated with terms elapsed after the surgery of congenital heart defects in children.

There are many data worldwide which suggest that the methods for evaluation of influenza vaccines immunogenicity should be improved. The only method validated in Russia is a HAI (haemagglutination inhibition) assay with serum samples from vaccinated volunteers. This assay does not, however, completely reflect the vaccine-induced immunological changes. In this study, we evaluated antibody immune responses to A (H5N1) inactivated influenza vaccine boosting in healthy volunteers previously primed with A (H5N2) live attenuated influenza vaccine. We compared three methods of antibody detection: (i) HAI assay with serum samples; (ii) ELISA with serum samples; (iii) ELISA with PBMC (peripheral blood mononuclear cells) culture supernates, i.e., an alternative test based on quantification of antibodies secreted by PBMC in vitro. The latter test was shown to have an advantage over other techniques in IgA and IgG antibody detection at early timepoints (day 7) after vaccination. The first two methods allowed immunogenicity assessment at day 28 after vaccination.

Thus, a test based on antibody quantification in PBMC supernatant samples can be used as an alternative method for evaluation of influenza vaccines immunogenicity. This method also exhibits a better strainspecificity.

The DR3 «death receptor» plays an important role in the initiation of apoptosis, proliferation, or inflammation. This receptor is shown to be involved in various diseases, including infectious conditions. Different variants of mRNA DR3 are formed as a result of alternative splicing. These variant transcripts encode membrane and soluble forms of the receptor which have different functions. Features of their expression and contribution of individual DR3 variants to the immune pathogenesis of infectious mononucleosis (IM) are poorely understood.

The purpose of this work was to develop, validate and test the techniques of DR3 gene expression assays, as well as to evaluate the DR3 mRNA splice variants by means of real-time RT-PCR and RT-PCR in the IM patients.

The original version of real-time RT-PCR allowed to determine relative amounts of DR3 mRNA, DR3 membrane variants (LARD1a + LARD8), and ratios of mRNAs encoding membrane and soluble forms of the receptor. The technique proved to be specific and sensitive (a semi-quantitative detection limit = 34-35 cycles) when tested in healthy volunteers and patients with acute infectious mononucleosis (AIM). Lower expression levels were shown for two alternative membrane variants of DR3 mRNA (LARD1b and DR3beta) thus regarding these isoforms as minor fractions. The relative levels of total DR3 mRNA expression were decreased in patients with AIM, as compared to healthy volunteers, whereas mRNA expression of membrane receptor variants did not differ between IM and controls.

To determine a qualitative contribution of either LARD1a and LARD8 variants into the expression of membrane forms of DR3, a two-step «nested» version of RT-PCR has been developed. It was shown that, in majority of control and IM samples, both main LARD1a, and alternative LARD8 membrane forms are contributing to mRNA expression of membrane DR3 variants.

The presented methods for evaluation of expression and occurrence of DR3 mRNA variants allow to characterize changes in the expression and function of this receptor associated with infectious mononucleosis. Further investigations are required to test the technique in larger number of samples.

MICA is a stress-induced protein that, as a rule, is not expressed in healthy tissues, but appears in large amounts on the surface of cells undergoing malignant transformation. In humans, this protein can either initiate antitumor immune response, or facilitate tumor cells for their escape of destruction. After shedding from tumor cell surface, soluble MICA enters blood circulation and contributes to decreased activity of effector cells, due to inactivation of NKG2D receptor. The aim of our study was to determine differences in circulating sMICA concentrations in sera of patients with different types of malignant lymphomas, and to evaluate the impact of sMICA upon NKG2D-positive cytotoxic lymphocytes. In experimental models with C1R-MICA cells, the MICA shedding was shown to occur from the surface of cultured tumor cells into the extracellular space. A reduced NKG2D expression dependent on sMICA concentration in the culture medium was demonstrated by flow cytometry in peripheral mononuclear cells, thus suggesting a role of serum sMICA in suppression of antitumor immune response. The sMICA detection was performed in patients with various types of B- or T-cell non-Hodgkin’s lymphomas by means of enzyme-linked immunosorbent assay. The groups of patients with increased sMICA content were identified and compared with the control group. Minimal amounts of serum sMICA were registered in the control group, with the median of 20 pg/ml. In a combined group of patients with various B-cell lymphomas, an increase in circulating sMICA amounts was shown, at the levels of more than 6.5 times exceeding the control values. The highest values of sMICA were recorded among the patients with chronic lymphocytic leukemia, diffuse large cell lymphoma, and multiple myeloma. Maximal sMICA levels among the investigated groups of patients were observed in the group of patients with T-cell anaplastic lymphoma (median of 574 pg/ml). The study provides preliminary evidence for a suppressive effect of different (immuno)chemotherapy components, in particular, rituximab and radiation therapy, upon serum sMICA contents in the lymphoma patients. Thus, elevated serum sMICA levels in patients with hematological malignancies may be considered as an additional criterion for application of the antitumor therapy. sMICA monitoring at different stages of cytostatic treatment may be useful in order to evaluate its efficiency.

Specific perinatal infections make about 30% in causal structure of infant mortality. Among the respiratory diseases of perinatal period, pneumonias take a special place, due to higher frequency, severity, complications and adverse outcomes. The aim of this work was to study microflora and factors of innate immunity (TLR2, TLR4, HBD-1, HBD-2, TNFα and NF-kB) at the level of the mucous membranes of the upper respiratory tract during intrauterine infection of fetus and perinatal pneumonia. Causal structure of ventilator-associated pneumonias at the intensive care unit represents a broad spectrum of pathogens with high resistance to antibiotics. Changes of immunological parameters (recognizing structures, i.e., TLR2, TLR4, HBD-1; HBD-2 defensins; proinflammatory TNFα cytokine and NF-kB transcription factor) in patients with intrauterine infections and pneumonia are ambiguous. Decreased expression of TLR2, TLR4 genes, along with increased of TNFα and NF-kB gene expression. These changes correlate with type of infectious pathogen, thus allowing us to assume that the pathogen, due to pathogenicity factors, may directly affect innate immunity mechanisms.

The study included thirty-one patients with coronary heart disease before the procedure of percutaneous transluminal coronary angioplasty with stenting and the day after the surgery. Flow cytometric assays were performed in order to determine major subpopulations of peripheral lymphocytes (CD3+CD19- , CD3- CD19+, CD3- CD16+CD56+, CD3+CD16+CD56+). A statistically significant decrease in the relative amount of total T-lymphocytes and increased relative content of NK and NK-T-cells was revealed in the patients under study when compared with healthy donors. The decrease in CD3+CD19- subpopulation did correlate with increased CD3- CD16+CD56+ lymphocyte set. We have not found any significant changes of immune profile at the early terms post-surgery, when compared with appropriate results obtained pre-surgery.

It has been shown that the influence of hormonal and non-hormonal therapy of menopausal syndrome upon immune profile may be variable, and the results of appropriate studies may be often contradictory. The aim of this work was to investigate the influence of hormonal and non-hormonal therapy in menopausal syndrome upon serum levels of some immunoregulatory proteins, i.e., alpha2-macroglobulin (a2-MG), pregnancy-associated alpha 2-glycoprotein (PAG), contents of some related cytokines (IL-6, IL-8, TNFα, IFNγ, IL-2) as well as VEGF amounts. Administration of menopausal hormone treatment and nonhormonal therapy was associated with reduced IL-6 levels in serum, regardless of treatment duration, or type of drug applied. We have found a statistically significant decrease of IL-8 serum levels in the course of dynamic monitoring in the women taking a menopausal hormone preparation containing 1 mg of 17β-estradiol and 5 mg dydrogesterone, and a non-hormonal drug containing genistein (60 mg) for 3-6 months. The levels of VEGF demonstrated high individual variability during therapy of menopausal syndrome. Serum concentrations of immunoregulatory a2-MG were stable in climacteric syndrome, and did not differ from normal values. However, the content of PAG, a known immunosuppressive protein, was increased 3-4 times in serum of 33-50% of the women receiving menopausal hormonal therapy, regardless of progestogen dose (5 or 10 mg dydrogesterone), and duration of its use. These findings suggest a need for individualized drug selection in order to minimize a risk of immunodeficiency conditions in the patients receiving hormone therapy of menopausal syndrome.

The aim of present study was to evaluate a possible role of Id2 (inhibitor of DNA-binding 2) in pathogenesis of bronchial asthma (BA). We have examined six healthy control persons, nine patients with allergic bronchial asthma (ABA) and twenty-four patients with non-allergic bronchial asthma (NABA). The Id2 mRNA expression was evaluated by means of real-time PCR.

No significant differences in Id2 mRNA expression levels were revealed between the control and asthma groups. Correlation analysis of Id2 mRNA expression has revealed significant positive correlation with AID (activation-induced cytidine deaminase) mRNA levels and peripheric blood eosinophil contents in ABA. Healthy persons demonstrate strong positive correlation between Id2 mRNA expression and serum IgE levels, like as with ε-chain mRNA contents.

It was suggested that the Id2 dysfunction may take place in the BA patients. These changes affect negative control over the AID expression, which is important for switching B-cells to IgE production.

Metabolic syndrome is a chronic subclinical inflammation. Published data on the immune state in metabolic syndrome are inconsistent, and, moreover, the gender features of metabolic syndrome are not described in details. The purpose of the study was to determine subpopulation profile of peripheral lymphocytes in young men with metabolic syndrome, as well as in cases of combined abdominal obesity and hypertension. The study included seventy-seven men aged 20-45 years. The men were divided into four groups matched by age: Group 1 represented healthy controls; group 2 included male patients with isolated abdominal obesity; group 3 consisted of males with a combination of abdominal obesity and hypertension; group 4, males with metabolic syndrome. The major lymphocyte subpopulations were isolated and analyzed by flow cytometry, i.e. T lymphocytes, T helpers, T-cytotoxic cells, TNK-lymphocytes, NK-cells, B-cells, as well as CD3+CD25+ and CD3+HLADR+ lymphocytes. In male patients with isolated abdominal obesity, we revealed increased relative contents of T lymphocytes, and increased proportion of the cells with CD3+CD8+, CD3+CD25+ phenotypes and higher B-cell numbers. A combination of abdominal obesity with hypertension in men was characterized by decreased ratio of T-helper/T-cytotoxic lymphocytes, due to reduced relative contents of CD3+CD4+ cells and increase in CD3+CD8+ lymphocytes. The numbers of T-helper cells in this group were significantly lower than in controls and among patients with isolated abdominal obesity. The number of T-cells with CD3+HLADR+ phenotype proved to be 2-fold higher than in the group of healthy male controls.