This review article concerns a challenging problem of HIV infection, immune activation. The processes of immune activation largely determine CD4+T cell depletion, leading to development of AIDS-associated and non-AIDS-related diseases. Immune activation indices are also strong predictors of the clinical outcome. Activation in HIV-infection affects both innate and adaptive immune cells, leads to increased proinflammatory cytokine production and blood coagulation. The reasons for immune activation may include different pathogens (HIV, cytomegalovirus, Epstein–Barr virus, hepatitis C virus), lymphopenia and lymphopenia-induced T lymphocyte homeostatic proliferation, transfer of microbial products from the gut, due to profound CD4+T cell deficiency in lamina propria and altered permeability of intestinal barrier. Other factors that promote the process of immune activation are as follows: T lymphocyte "bystander" activation, increased T cell turnover, and systemic inflammation development. The review covers pathogenic mechanisms HIV-infection associated with immune activation, like as description of different laboratory parameters characterizing its manifestations in HIV-infected patients. Significance and prognostic role of these parameters in assessing efficiency of antiretroviral therapy, development of complications, and adverse outcomes of infection are presented as well.

Neonatal screening for primary immunodeficiencies (PID) provides a unique opportunity for early detection of moderate to severe immune disorders. Thus, the patients could be provided with best available therapy, due to effective treatment according to updated PID management guidelines. Clinical outcomes of recently implemented screening programs have demonstrated high sensitivity and specificity of the diagnostic techniques used and good survival rates. The existing evidence provides a strong basis for inclusion of PID screening into general Russian neonatal screening program.

The safety issues of human intravenous immunoglobulin preparations are particularly important in modern pharmacotherapy for immunodeficiencies, hematologic and neurologic diseases, like as at transplant centers. Upon massive infusions of these media some complications are detected that are associated with spontaneous activation of complement system accompanied by production of anaphylatoxins, as well as activation of kallikrein/kinin, plasmin, and blood coagulation systems, changed blood rheology, initiation of intravascular hemolysis. For distinct groups of patients, these complications may be due to presence of some anti-erythrocyte antibodies (e.g., anti-A and anti-B haemagglutinins, anti-D antibodies) in the intravenous human immunoglobulin preparations. In the present review article, we show development of current quality standards for human intravenous immunoglobulins based on determination of antibody contents. Antibodies to erythrocytes represent a special safety index aiming to minimize risk of possible adverse effects connected with transfusions of human blood preparations. Different haemagglutination tests were compared to assess contents of anti-A, anti-B haemagglutinins and anti-D antibodies for specific safety of human intravenous immunoglobulins. Analysis of haemagglutination techniques for evaluation of human intravenous immunoglobulin preparations revealed their relative advantages and disadvantages. Various modifications of the methods are discussed, thus allowing to optimize process of quality control for these preparations based on detection of haemagglutinins and anti-D antibodies. We demonstrate a necessity to adjust regulations and to improve evaluation techniques for haemagglutinin determination in human immunoglobulin preparations at amounts of 100 mg/ml of protein. Special features of Russian national quality standards for human immunoglobulin preparations are considered with respect to assessment of haemagglutinins and anti-D contents. One may conclude that haemagglutination methods present the most informative and economically substantiated approach when assessing specific safety of human intravenous immunoglobulins by measuring contents of anti-A, anti-B haemagglutinins, and anti-D antibodies.

There is a growing body of data about the cytopathic effect of bacterial arginine deiminase on human endothelial cells, but the precise mechanisms of endothelial dysfunction caused by the activity of the enzyme remain poorly understood. Activity of arginine deiminase causes arginine depletion in the microenvironment of the host organism cells. In view that arginylation of beta-actin regulates actin cytoskeleton structure and cell motility, we proposed that the cytopathic effect of arginine deiminase may be associated with disruption of actin in the cytoskeleton of endothelial cells. The aim of this study was to investigate the effect of arginine deiminase from S. pyogenes on migration and actin cytoskeleton structure of the human endothelial cells, line EA.hy926. The supernatant of sonicated S. pyogenes M49-16, its isogenic mutant with a deletion of the arginine deiminase gene (S. pyogenes M49-16delAD), supernatant of sonicated S. pyogenes M22, and arginine deiminase isolated from the latter strain were used. The effect of bacterial factors on migration activity of endothelial cells was studied in the model of "wound healing" in vitro. To analyze the influence of bacterial factors on the actin cytoskeleton structure, cells were stained with phalloidin-rhodamine. It was shown that supernatants of destroyed S. pyogenes, as well as arginine deiminase significantly reduced the migration activity of endothelial cells and altered the structure of their actin cytoskeleton. The supernatants of destroyed S. pyogenes M49-16delAD with  deleted gene of arginine deiminase showed a significantly reduced ability to suppress cell migration as compared with the supernatant of sonicated S. pyogenes M49-16. No significant differences were revealed in the structure of actin filaments in cells cultured in the presence of supernatants of destroyed S. pyogenes M19-16, and cells cultured in the presence of isogenic mutant S. pyogenes M4916delAD. Adding exogenous arginine to the cells cultured with supernatants of destroyed S. pyogenes did not restore their migratory activity and the structure of their actin cytoskeleton. However, if arginine deficiency caused by the activity of arginine deiminase was compensated, endothelial cells migration activity was restored, and the structure of actin cytoskeleton was recovered. A decrease of migration activity of endothelial cells under the influence of streptococcal arginine deiminase was due to the disruption of actin cytoskeleton structure.

CD3+T lymphocytes were isolated from normal donors by positive magnetic separation. Activation of the T cells with particles conjugated with antibodies to CD3, СD28 and СD2 molecules led to substantial increase in T cell production of interleukin-8 (IL-8). An interleukin-8 receptor (CXCR1, CD181) was initially expressed in 13.3% of T lymphocytes. Activation of T lymphocytes resulted into a detectable increase of CD181+ cell number among CD4+ naïve cells and CD4+ terminally-differentiated effector cells, and, conversely, into decrease of their number among CD4+ effector memory cells. Activation of T lymphocytes was assessed by membrane expression of CD25 molecule (receptor for IL-2). IL-8 (0.01-10.0 ng/ml) was shown to markedly reduce activation of both CD4- and CD4+ effector memory T cells, as well as terminallydifferentiated T effectors, without significantly affecting activation of naive T lymphocytes and central memory T cells. IL-8 noticeably increased IL-2 production by activated Т cells, caused a reduced IL-10 production, and did not significantly affect the secretion of IFNγ and IL-4. The data obtained suggest a significance of IL-8 for direct regulation of adaptive T cell responses.

Age-related macular degeneration is one of the most widespread multifactorial eye diseases. Polymorphic functional alleles of vascular endothelial growth factor (VEGF) combined with matrix metalloproteinase (MMP) gene and tumor necrosis factor (TNF) gene variants may influence the development of disease. We have performed frequency analysis of their polymorphisms in regulatory regions of VEGF (rs 699947, rs 3025039), ММР2 (rs 2438650), ММР3 (rs 3025058), ММР9 (rs 3918242), TNFα (rs1800630, rs1800629, rs 361525) genes, and their combinations in a group of patients with age-related macular degeneration (MD). Frequencies of TNFα (rs1800629) genotypes significantly differed for the MD patients and control group. Upon the combined genotype analysis, we have revealed six constellations of VEGF-ММР genes that were positively associated with the disease development. Five of them included minor homozygous genotype VEGF-2578АА. A combined analysis of VEGF – TNFα genes polymorphisms has shown presence of both positive and negative complex genotypes. The most significant differences have been detected by comparative analysis of the complex genotypes frequencies which included 8 polymorphic regulatory gene regions of all genes studied. In most genetic complexes associated with the disease development, homozygous TNFα-863СС, homozygous MMP2-1306 ТТ, and MMP9-1562СС genotypes have been detected, together with the combination of homozygous VEGFA+936СС genotype in the same patients. We can assume that harboring allelic variants, which may contribute to angiogenesis prorcesses is typical for the genome of patients with macular degeneration, along with low-level production of pro-inflammatory regulatory factors and enzymes participating in degradation of extracellular matrix. Analysis of complex genetic factors, procing some factors taking part at the pathological process being the regulators of production for each other, is more informative when detecting protective and resistant markers of the disease development rather than single genetic markers, thus being useful for genomic screening.

The objective of our study was to evaluate production of matrix metalloproteinase–2 and -9 in  lacrimal fluid, as well as СFH gene polymorphism in a group of patients with primary open-angle glaucoma.  The study was performed with 120 eyes of patients with primary open-angle glaucoma (POAG) including  34 eyes from the first-stage disease; 35 eyes, from the 2nd stage; 35, at the 3rd stage of the disorder, and 16  eyes with stage 4 POAG. Control group consisted of 25 glaucoma-free patients. Tear fluid served as a material  for biochemical tests. The levels of MMP-2 and MMP-9 were determined by sandwich ELISA for the both  eyes. Venous blood was used for genetic studies. Genomic human DNA was isolated from the whole blood  leukocytes, a polymerase chain reaction (PCR) was performed.  The results of this study were analyzed using a package of SAS applied statistical programs (Statistical  Analysis System, SAS Institute Inc., USA) employing standard algorithms of variation statistics, i.e., correlation  analysis, contingency tables, and various types of intergroup comparisons for the parameters studied. The  following results were obtained: average indices for metalloproteinases-2 and -9 were found to be increased upon  progression of the disease. Correlation analysis of contingency relationships was performed for morphometric,  clinical, functional parameters, and quantitative indices of MMP-2 and MMP-9. We also studied prevalence  of CFH gene polymorphismic alleles depending on the stage of glaucoma development. One may conclude  that our study provided evidence for a relationship between MMP-2 and MMP-9 production, and СFH  gene polymorphism, thus expanding a list of host genetic mutations which could be potentially involved into  pathogenesis of primary open-angle glaucoma.

We searched for a possible target tissue in eye orbit for thyroid autoantibodies in endocrine ophthalmopathy (Graves’ disease), using correlation analysis method. We examined a group of 139 patients (278 eye orbits) with thyroid-associated ophthalmopathy associated with diffuse toxic goiter. Serological parameters (antibodies to thyroid-stimulating hormone receptor; thyroglobulin, thyroid peroxidase) were compared with instrumental diagnostic data (multi-layer CT, ultrasonography of eye orbit, and exophthalmometer), as well as clinical symptoms. Statistical correlation analysis enabled us to show different degrees of association between thyroid antibodies and clinical manifestations of Graves’ disease and eye orbit involvement. Especially, carriers of antibodies to TSH receptor and thyroglobulin (as compared to seronegative patients) exhibited higher exophthalmos scores (19.16±0.26 mm, p < 0.001, and 19.41±0.40 mm, p < 0.05, respectively), and with total muscle index (2.42±0.05, p < 0.01, and 2.42±0.08, respectively). Meanwhile, eyelids in carriers of antibodies to TSH receptor and thyroid peroxidase proved to be more swollen (p < 0.001, p < 0.05, respectively). Carriage of antibodies to thyroglobulin was associated with synchronous involvement of two structures of the eye orbit: extraocular muscles and retrobulbar tissue, which is reflected by increase in the average ntegral exophthalmos index within the group.

It is known that a relationship exists between patient’s age and pathogenesis of malignant tumors associated with ability of tumor and its microenvironment to secrete cytokines that may play an important role in carcinogenesis. The aim of present study was a comparative analysis of cytokine production by peripheral blood cells and tumor samples when exposed to polyclonal activators (PA) in patients of different age groups with invasive ductal carcinoma. Peripheral blood samples of 54 women with invasive ductal carcinoma were under study. The patients were divided into the following groups: I, 40-60 years old; II, 61 year and older. Metastases into lymph nodes were revealed at the moment of study in 13 patients (42%) from group I. By contrast, only 3 of 23 (10%) patients with metastases were revealed in group II. Fisher’s ratio test showed significant differences between occurrence of metastases in lymph nodes, and patient age (p = 0.034). In both groups of patients, there was an increase of IL-6, IL-8, IL-10, IL-17, IL-18, IL-1ra, G-CSF, GM-CSF and VEGF concentrations in tumor supernates and its microenvironment, when compared with the production of cytokines by blood cells, thus indicating to high functional activity of tumor together with its microenvironment. Effect of PA led to increase of IL-2, IL-10, IL-17, IL-1β, IL-1ra, TNFα and IFNγ concentrations in blood cells supernatant which suggested their high ability to produce cytokines. In the group of patients who were 40-60 years old, the stimulatory index of PA (SIPA) for IFNγ production was several times higher than similar index in patients aged 61 years or more. Increase of SIPA for all cytokines in blood cells supernates, in comparison to SIPA of tumor supernates, could be explained by initially high levels of spontaneous cytokine production by the tumor and its microenvironment that are able to promote tumor growth. Therefore, effect of PA upon tumor tissue was not so expressed as its effect upon immunocompetent blood cells. Both tumor and microenvironment maintained high ability of IL-18 production, thus confirming its important role in carcinogenesis. In summary, these findings suggest that circulating immunocompetent cells may serve as an additional factor providing tumor progression, due to their high ability to secrete cytokines.

Antibodies against sex steroid hormones are known to modulate their serum concentration and  inhibit experimental breast cancer (BC). Hence, these effects could be changed by according anti-idiotypic  antibodies. However, relationships between antibodies and anti-idiotypic antibodies and BC in women are still  poorly studied. The aim of this study was to identify possible associations between occurrence of antibodies  against estradiol and progesterone (IgG-Es1and IgG-Pg1) and according antiidiotypic antibodies (IgG-Es2  and IgG-Pg2), and postmenopausal BC.  Eighty-nine  healthy  women  and  273  BC  patients  were  examined.  A  non-competitive  solid  phase  immunoassay for IgG-Es1 and IgG-Pg1 was performed using estradiol and progesterone conjugates with bovin  serum albumin as antigens. Monoclonal antibodies against Es and Pg as antigens have been used for noncompetitive solid phase immunoassay of IgG-Es2 and IgG-Pg2.  Results: absence of both IgG-Es1 and IgG-Pg1 was revealed in 53.9% of healthy donors and 41.0% of  BC patients (p = 0.04; OR = 0.6). Presence of IgG-Es1 without IgG-Pg1, or IgG-Pg1 without IgG-Es1was  detected for, respectively, 15.7% and 20.2% of healthy women, and in 10.3% and 7.7% of BC patients (p > 0.05).  Simultaneous increase of both IgG-Es1 and IgG-Pg1 was revealed in 10.1% of healthy donors and in 41.0%  of BC patients (p < 0.0001; OR = 5.3). Absence of a single antibogy (IgG-Es2 or IgG-Pg2) showed similar  frequency in the groups under study. Simultaneous increase of both IgG-Es2 and IgG-Pg2 was detected in  52.8% of healthy women and 34.8% of BC patients (p = 0.03; OR = 0.4). Conclusion: We have first revealed  an immunostimulating synergistic effect of antibodies against sex steroid hormones and immunoinhibitory  synergistic effect of appropriate anti-idiotypic antibodies upon the postmenopausal breast cancer risk.

Associations between IL1B (rs1143634), IL1RN (VNTR, intron 2), IL4 (VNTR, intron 3), TNFA (rs1800629, rs361525), IL6 (rs1800795), and IL10 (rs1800896) genetic polymorphisms in women with recurrent miscarriage (RM) were analyzed. We studied DNA samples of 112 women with RM and 267 women with physiological pregnancy. The IL1RN, IL4 genotypes were identified by PCR techniques, the IL1B, IL6 gene polymorphisms were defined by means of RFLP approach. To detect TNFA and IL10 gene polymorphisms, TaqMan real-time PCR was used. The results have shown that polymorphic loci of IL1B, IL1RN, IL10, TNFA genes were not associated with RM, and early spontaneous abortion risk. The 2R allele of IL4 gene was found to be associated with higher RM risk (OR = 1.52; 95% CI = [1.08-2.14]; P-value (cor) = 0.05), and G allele of IL6 gene was associated with a risk for > 3 early spontaneous abortions (OR = 2.10; 95% CI = [1.24-3.56]; P-value (cor) = 0.05), in an additive inheritance model. Upon evaluation of the data obtained, one may conclude that the IL4 (VNTR intron 3) and IL6 (rs1800795) gene polymorphisms could influence the RM development. These results may be useful for assessment of molecular mechanisms underlying early spontaneous abortion.

The aim of the work was to study the parameters of T-cells, humoral immunity and activity of NAD(P)- dependent dehydrogenases in lymphocytes from peripheral blood at young children with recurrent obstructive bronchitis (ROB). Patients and Methods: Fifty-four children at the age of 1-3 years with ROB were observed. The control group consisted of 35 healthy children at similar age. The numbers of СD3+, СD4+, СD8+, СD19+ cells in peripheral blood were determined by flow cytofluorimetric technique. IgA, IgM, and IgG concentrations in blood serum were evaluated by G. Mancini et al. (1965); circulating immune complexes (CIC), by V. Haskova et al. (1978); total IgE test was performed with “Total IgE – IFA Best” system (Russia). Relative activity of NAD (P)-dependent dehydrogenase in peripheral blood lymphocytes was studied according to A. Savchenko et al.(1989). The results of this study have revealed changed phenotypic spectrum of peripheral blood lymphocyte populations; a statistically significant reduction in IgA concentration, and a tendency for increased IgE levels in blood serum in the children with ROB. An increase of ribose-5-phosphate- and NADPdependent metabolic processes; reduced lactate dehydrogenase anaerobic reactions; a switch from lipid catabolism products to glycolysis; a shunt role of malate/aspartate in cell energy supply, and for glutathione reductase activity; elevated levels of substrate flow along Krebs cycle in peripheral blood lymphocytes were registered as well. It may be concluded that the change of phenotype and enzymatic activity of blood lymphocytes, selective IgA deficiency in blood serum could be shown in the younger children with recurrent obstructive bronchitis.

Atopic bronchial asthma (ABA) is a multifactorial disease; its development is dependent on many environmental and genetic factors. Genetic risk factors can affect the clinical phenotype of ABA and the level of therapeutic control over the disease. Cytokine genes are crucially important in pathogenesis of ABA as they encode proteins participating in immune response and development of inflammation in bronchi. It was suggested that the therapeutic control of the disease is genetically mediated and depends on the presence of one or another allele in genes of mediators, participating in ABA pathogenesis. The knowledge about genetic markers will allow to predict clinical course of ABA in children. We carried out the analysis of association between genes of pro- and anti-inflammatory cytokines with the level of therapeutic control of ABA. In children with controlled and uncontrolled ABA (CABA and UABA, respectively; n = 110), and in general a population sample (n = 138), we analysed 11 polymorphisms: IL2 (rs2069762), IL4 (rs2070874 и rs2243250), IL5 (rs2069812), IL10 (rs1800872 and rs1800896), IL12B (rs3212227), TNFA (rs1800629 and rs1800630), TGFB1 (rs1800469), and IFNG (rs2069705), encoding cytokines actively participating at the development of allergic inflammation. According to results of present study, the prevalence of alleles and genotypes of the analysed loci in the East Siberia Caucasians is consistent with the data in other world Caucasian populations. We have found statistically significant differences between UABA and control groups for the prevalence of IL2 (rs2069762) polymorphism: GG genotype was more common in control group (14.1% compared to 5.9%, р = 0.03). It was shown that the IL2*T allele and ТТ genotype of the rs2069762 are associated with the increased risk of uncontrolled ABA. A comparison of the haplotypes of IL4 (rs2070874 and rs2243250) gene with correction for sex and age within an additive model revealed that the most common haplotype СC (prevalence in ABA/CABA/UABA groups is 0.75/0.76/0.74, respectively) is protective against the development of ABA (RR 0.53±0.32; p = 0.044). A comparative analysis of TNFA (rs1800629 and rs1800630) haplotypes has shown the GC haplotype to be protective against the risk of ABA (RR 0.59±0.17; р = 0.003) while the GA haplotype is positively associated with the disease (RR 2.07±0.25; р = 0.003). This was true for BA, regardless of the control over the disease (CABA GA: RR 1.93±1.2; p = 0.041; UABA GA: RR 2.43±0.31; p = 0.005). Thus, it was established that the studied cytokine genes are important for the development of ABA in children. These data were obtained for the first time for Caucasians of East Siberia. They are of interest in terms of accumulation of the data about the impact of cytokine genes polymorphism upon development of ABA and its therapeutic control in children.

The article considers the experience of Russian and leading foreign regulatory agencies in organisation and conduction of preclinical and clinical trials of human immunoglobulin products. The authors suggest a classification of human immunoglobulins and provide updated information on authorization of these products in Russia. The article summarizes methodological approaches, basic scientific principles and criteria relating to expert evaluation of preclinical and clinical trials of blood products. The authors further define the expert body’s requirements for data on preclinical and clinical trials of human normal immuniglobulins and human specific immunoglobulins for the prevention and/or treatment of infectious and non-infectious diseases which are submitted as part of applications for marketing authorization or marketing authorization variation. The article suggests programs of preclinical and clinical trials for human normal immunoglobulins and human specific immunoglobulins for the prevention and/or treatment of infectious and non-infectious diseases that are aligned with the Russian legislation and Eurasian Economic Union’s regulations on medicines circulation, and have been elaborated with respect to the guidelines of the European Medicines Agency.

We have investigated in vivo effects of quercetin dihydrate (QD) upon course of respiratory allergy produced in experimental murine model. To provoke allergy, serum ovalbumin (OVA) was injected at different terms. Following administration of QD at the stage of OVA sensitization, the balance of T-helper cytokines was changed (IL-2 was decreased along with increased IFNγ) accompanied by decreased allergen-specific IgG1, IgE and increased IgG2a in blood serum. At the stage of boosting the respiratory allergy, intravenous administration of QD caused reduction of inflammatory lung infiltration, and composition of bronchoalveolar lavage fluid was altered, i.e., IgG1 and IgE showed a decrease, along with higher IgG2a levels.

Type 1 diabetic mellitus (T1DM) is known to be associated with progressive destruction of β-cells of the pancreas. Dysregulated immunity and programmed cell death are an important link in pathogenesis of diabetes. In this study, we examined expression levels of interleukin-2, Bcl-1, ANXA-11 genes in patients with T1DM. The study was done with blood leukocures of T1DM patients (30) and healthy controls (70). Reverse-transcription PCR was done with Transcriptor First Strand cDNA Synthesis Kit from Roche Life Science. The study of IL-2 and Bcl-1 gene expression level in peripheral blood leukocytes indicated that the median gene expression levels of IL-2 and Bcl-1 are increased in patients with T1DM patients compared with control group of healthy persons. The study of apoptosis by annexin test has revealed an increased level of ANXA 11 expression in T1DM patients. The obtained data can serve as an additional source for understanding the pathogenesis of T1DM mechanisms.

Chronic gastritis is the most common disease of gastro-intestinal tract. Precancerous potential is among most important epidemiological features of chronic gastritis. Immune system plays a distinct role in transformation from precancerous state to malignancy. In this context, the aim of our work was a study of spontaneous and induced chemiluminescence activity of neutrophilic granulocytes in patients with chronic superficial gastritis, chronic atrophic gastritis and gastric cancer. The work presents results of comprehensive laboratory examination of patients with chronic gastritis (CG) (a total of 85 persons. 25 patients with chronic atrophic gastritis (CAG), and 50 patients with gastric cancer (GC) at the age of 19 to 70 years were enrolled. Control group included 115 healthy donors without gastrointestinal complaints at the age of 19 to 67 years. The study was performed with venous blood samples taken from cubital vein into Vacutainer tubes with sodium heparin (5 U/mL) prior to starting any pathogenic treatment. Evaluation of spontaneous and induced chemiluminescence was performed for 90 minutes at a 36-channel “CL 3606” chemiluminescence analyzer (Russia). In our study, patients with gastric cancer showed clear unidirectional changes in chemiluminescent activity of neutrophilic granulocytes (NG). When measuring spontaneous and induced NG chemiluminescence, we diagnosed a decreased phagocytic activity characterized by prolonged time-to-peak and area under the curve for spontaneous and induced CL, thus presuming longer activation time required in cases of reduced phagocytic function. The NG activity in patients with chronic gastritis is not impaired, but, similar changes of time-to-peak and area under were detected. Chemiluminescent activity of NG is increased in the group of CAG patients, and, considering similar changes in activation time and area under the curve, NG also produce greater amount of reactive oxygen species. Thus, for all H.pylori-associated diseases, the blood NG take longer time to activate, whereas their functional activity is not disturbed in CG, enhanced in CAG patients, and reduced in gastric cancer.