[REDUCED RESPONSE OF NATURAL KILLER LYMPHOCYTES TO TOLL-LIKE RECEPTOR 3 STIMULATION IN CHILDREN WITH RECURRENT INFECTIONS.]

In this study, we examined the activation of natural killer(NK)-lymphocytes mediated by Toll-like receptor 3(TLR3), in the group of children with recurrent infections and the group of children with invasive bacterial infections. We examined level of CD69 (marker, of activation) expression on NK - lymphocytes after incubation with TLR3 ligand. There was a significant decrease in the level of an activation marker - 36,3±4,4% in the group of children with recurrent infections, compared with a control group of healthy children (56,5±4,9%) and the group of children with invasive bacterial infections (55,9±4,4%). Also, decreased was an activation potential of NK-lymphocytes - 24,9±L4,5% which was calculated as the difference between the percentage of CD69+ NK - lymphocytes after incubation and spontaneous value (without the addition of activator). There were no correlations between markers of TLR3 mediated activation of NK-cells and the age. Thus, the ability to activate NK - lymphocyte mediated by TLR3, independent of age and reduced in the group of children with recurrent infections. Reduced activation of NK-lymphocytes may contribute to increased susceptibility to viral infections and bacterial complications.

[Effect of hypo- and hyper- accentuated NK cell activity on embryo implantation].

NK lymphocytes play an important role in implantation and during development in early pregnancy. Recently, we showed that the proportion of NK that expressed CD69 after incubation with K562 (CD69(stim)) cells reflected the NK population excitation potential. In the present study, we investigated the significance of NK activation levels in predicting the implantation outcome in 84 patients following IVF (in vitro fertilization). Remarkably, the patients with an accentuated increase or a decrease of the levels of NK activity, have unfavourable conditions for implantation (9.1%, 3/13 and 15.1%, 5/25, respectively) compared to the patients with the nominally normal levels (52.2%. 25/46). Therefore, a nominally normal level of the NK activity is an important physiological condition and predictive factor for immune readiness to embryo implantation. This study describes an easy, efficient, sensitive and informative method for measuring NK cell activity that is relevant to clinical trials.

[Qualitative analysis of accented CD158a receptor expression in NK-lymphocytes in women with reproductive failures].

Blood from 91 women that undergoing IVF cycle was analysed for NK cells CD158a expression using a monoclonal antibody and FACScan flow cytometer and CellQuest software (BD Bioscience, San Jose, USA). Patients were separated on 3 groups according implantation and pregnancy results in actual IVF cycle. 53 patients not became pregnant (IVF failure group F), 24 became pregnant after IVF with subsequent successful pregnancy (Pregnancy succes group PS) and 13 became pregnant with subsequently pregnancy failure (Pregnancy failure group PF). Average levels of of CD158a on NK cells were significant increase in patients that not became pregnant compared to pregnant group. However IVF failure patients have comparable average CD158a levels to reproductive success group. Patients with pregnancy failure have significant decreased CD158a levels compared to both IVF failure and reproduct success patients. A qualitative analysis of NK CD158a expression showed that 22/24 (92.8%) women who became pregnant and live birth had CD158a levels that were > 20 but < 65%. In contrast only 62.8% patients form IVF failure and 61.6% from Pregnancy failure group had CD158a expression on NK in this zones (corridor). 38.4% of patients from pregnancy failure group had CD158a expression levels lower than 20% and as a result significant decreased average value in whole group. In contrast IVF failure patients had increased CD158a expression in 9.5% cases and decreased in 27.7% and as a result similar average levels to pregnancy success groups. Decreased CD158a expression (< 20%) was significant predictive factor for reproductive failure (OR 10,7) Increased CD158a expression > 65%) was predictive factor for Implantation failure (OR 5,4; P = 0,09) Normal CD158a expression (> 20% but < 65%) was significant predictive for IVF implantation and Pregnancy success and as a result for common Reproduct success (OR 2,7; 6,87; 6,92). We found that normal NK CD158a expression is associated with successful IVF and pregnancy. Preference of qualitative analysis under simple average value comparison in case of bilateral distribution of parameters was shown.

[Functional and morphologic changes in the tendons and multimicrovacuolar collagen absorbing system of the three-phalanx fingers in disorder of arterial inflow].

Taking into account the fact, that unsatisfactory results of treatment of the thumbs traumatic damage are significantly caused by presence of anatomic disruption of the thumbs palm's own arteries with chronic tissue ischemia occurrence. One of consequences of damage of the three-phalanx thumbs interphalanx joints and the thumbs palm's own arteries is a formation of quickly progressing secondary flexing contracture of proximal interphalanx joints (PIPHJ). In the investigations accomplished there was established, that the changed tissues of the flexors tendons and their sheaths, forming cicatricial blockade distally to the primary trauma site, are taking part in a PIPHJ contracture development while arterial perfusion presence.The tissues damage severity, as well as a severity of clinical features of the tendons and their sheaths damage, depends on a degree of an arterial perfusion disorder present,

[The immunophenotypic characteristics of two functionally different natural killer cell subpopulations in peripheral human blood].

NK cell cytotoxicity and immunophenotypic characteristics of activated natural killer (NK) cells after co-incubation with K562 target cells in women were investigated. An increase in CD69, HLADR, CD95 expression on target-activated NK cells was demonstrated. Conversely, CD62L expression in NK cells after activation was decreased. The results showed that only a part of NK cells after incubation with K562, became CD69 positive. Moreover, even after lowering of "efector - target" ratio and extensions of time of stimulation, the amount of CD69+NK cells did not exceed 60% (50% +/- 1.7%). Expression of CD69, CD95 and HLADR on NK cells before coincubation didn't correlate with NK cytotoxicity. However, size of population of activated CD69+ NK cells (after stimulation) was correlated with NK cytotoxicity. We also found that the activation capacity after cocultivation with K562 cells is related to loss in CD62L on NK cell's surface. Only CD62L(neg) and less CD62L(low)NK cells had the ability to be activated with K562. In conclusion, we demonstrated that incubation with K562 target cells enhanced the expression of CD69 on the major part of NK cell population and the size of CD69+NK cell population is strongly correlated with NK cytotoxicity.

[Experiment study of multimicrovacuolar absorbing collagenic system and its role in trophic processes of nonsynovial part of the tendons].

The arterial inflow disorder and occurrence of an acute and subsequent chronic ischemia constitutes frequent complication of a hand and fingers trauma. One of the consequences of ischemic damage of tissues is formation of a flexor contracture of fingers, involving the flexors tendons. In extrasheath zone the blood supply is secured by paratendon--a connective tissue, taking part in the tendons sliding. According to data, obtained during morphological investigation of paratendon in laboratory animals, there was established its principal similarity with multimicrovacuolar collagenic absorbing system of a man. She constitutes a spatial porous structure, packed with cellular elements and vessels. While simulation in experimental conditions of the blood flow disorders, which in physiological environment is secured by paratendon, alike in disorder of the bone-tendon and muscle-tendon pathways of blood supply, there were revealed the coarse changes in the tendon structure and pronounced adhesion of tendons with loss of their sliding capacity, what had permitted to consider the paratendon blood circulation a dominating one.

[Application of Rheosorbilactum in the therapy of traumatic shock: comparative study].

The authors have studied influence of infusion of 800 ml RHEOSORBILACTUM, 500 ml VENOFUNDIN and 800 ml of 0,9% isotonic solution of sodium chloride with speed of 50-55 ml/mines on hemostasis, the maintenance of a liquid in the thorax, hemodynamic parameters in 60 patients with traumatic shock. It was revealed that the use of investigated volume of RHEOSORBILACTUM increases reliably (P<0,05) oxygen delivery for the account of the expressed haemodynamic effect which duration is less than during the use of VENOFUNDIN. It was revealed also that RHEOSORBILACTUM increases reliably (P<0,05) quantity of liquid in the thorax without an considerable difference with other investigated medications. There was no influence of RHEOSORBILACTUM on time of trombus formation tests--prothrombin time, the international normalized index, activated partial thromboplastin time.

[Anti-idiotypic antibodies as a physiological mechanism of inhibition of cofactor-independent antiphospolipid antibody formation].

Antiphospholipid antibodies (APA) and anti-idiotypic antiphospholipid antibodies (AiAPA) were studied in 148 women subjected to IVF. APA (IgG aCL, aPS,) levels in serum have been defined. Sera IgG fraction was also examined for the presence of AiAPA by three different methods: 1) binding of AiAPA with mouse monoclonal cofactor-independent APA (mAPA) immobilized on plate; 2) AiAPA neutralization of human affinity isolated APA and 3) mAPA binding with phospholipids. Significant difference in AiAPA levels between APA+ and APA- women subjected to IVF have been found. The mean level of AiAPA was lower in APA women subjected to IVF than in APA- women from the same group (p < 0.05). IV infusion of Ig decreased the APA level significantly as well as increased the AiAPA level in APA+ women subjected to IVF. Ig application to incubation in vitro results in decrease of APA levels in serum. Results of this study confirmed possibility of AiAPA participation in down regulation of cofactor-independent APA production in women undergoing to IVF.

[Molecular diagnosis and frequencies of primary hypolactasia in populations of RUSSIA and neighboring countries].

The digestion of the milk sugar (lactose) is observed every normal child but not in every adult. The decreased lactase synthesis in some adults results in problems with digestion of the whole milk (primary hypolactasy). An association of lactase activity in adults with carrying of the allele T within the polymorphism C/T-13910 located upstream of the lactase gene and 100% association of hypolactasy with the genotype C/C has recently been shown for a Finnish sample. In the present work we determined the LCT* C/T_13910 genotypes and allele frequencies in populations from Russia. The genotype C/C frequencies varied from 36.6% for Russians to 88.2% for Chukchi and were close to the published medical and epidemiological data on hypolactasy frequencies in respective populations. Genotyping was performed by three different methods to identify the optimal one. Our results have shown that the studied locus is the key determinant for the primary hypolactasy development in various human populations. Consequently, the DNA diagnostics of the C/C genotype carrying is a promising predictive test to detect the primary hypolactasy long before its clinical development. Practical application of this type of diagnostics would be a step towards the individual-oriented medicine.

[Studying leucocyte-endothelial mechanisms for diagnosis of vascular disorders in vibration disease].

The article presents pathogenetic mechanisms of microangiopathy development in vibration disease. The authors specified leucocyte-endothelial interactions in vascular dysfunction caused by local vibrations, suggested additional methods to diagnose vascular disorders in vibration disease patients.

[Immunopathophysiologic characteristics of early pregnancy in women with recurrent miscarriage].

UNLABELLED: The lymphocyte subsets, activation marker expression and activity of ConA-treated lymphocytes have been studied in 58 patients with a history of unexplainable pregnancy loss (UPL) and 22 normal pregnant women (control) in 4-7 weeks of pregnancy. The increase of CD16/56+ cell level and CD4+/ CD8+ ratio and decrease of CD19+ cell level have been found in peripheral blood of UPL patients in comparison with control. The expression of HLA-DR was upregulated on CD3+ and CD8+ cells and the expression of CD25 was downregulated on CD3+, CD4+, CD8+, and CD16/56+ cells in UPL women. According to correlation analysis results, low expression of CD25 was related to a low expression of CD8 on NK, high expression of CD45RA on CD4+ helper cells, high expression of HLA-DR on CD8+ cytotoxic cells. High frequency of ConA-induced activation, low frequency of ConA-induced suppression and low suppressive activity of ConA-induced lymphocyte were found in UPL patients compared to control. CONCLUSION: women with UPL have disorders in feto-maternal recognition in early pregnancy that led to a development of the inadequate immune response to fetus realized as a defect of NK activation, deficiency of T-cytotoxic cell limitation and memory helper cell generation, downregulation of TR cells and suppressor function.

[Differential diagnosis of specific and nonspecific lung diseases by the parameters of the leukocytic enzymatic system].

The study examined 56 patients who were divided into three groups: 1) patients with infiltrative pulmonary tuberculosis; 2) those with nonspecific pneumonia; 3) control (apparently healthy individuals). In addition to the routine studies, the R. P. Nartsissov procedure was used to determine the activity of SDH and alpha-GPDH in all the examinees. The findings indicate that the lymphocytic activity of basic energy processes is much decreased in infiltrative tuberculosis and nonspecific pneumonia. Of the greatest value for active diagnosis is the determination of alpha-GPDH activity that is reduced only in patients with infiltrative pulmonary tuberculosis, its low activity points to profound redox disorders.

[Results of immunization with allogenic leukocytes in women with recurrent pregnancy loss].

Immunization with allogenic leukocytes has been proposed as immunotherapy of recurrent pregnancy loss. However efficacy, mechanism and immune monitoring of it remain not fully explainable. Our results suggest high clinical efficacy of alloimmunization related to normalizing effect on subsets, activation and suppressor function of lymphocytes, and placental protein production. Based on data obtained, the possible use of immune parameters and placental proteins for determination of alloimmunization efficacy was proposed.

[Analysis of specificity of antiphospholipid antibodies with the use of monoclonal antibodies to phospholipids].

The antigenic specificity of antiphospholipid antibodies (APA) is a matter of intensive investigation. Difference in the reported involvement of APA in clinical manifestation may be due, in part, to the polyclonal nature of these antibodies and the use of serum and serum fractions for analysis. To circumvent this issue, we generated mouse monoclonal APA and compared their antigen binding patterns and conditions of this reaction. Monoclonal APA 5A1 and 1B10 reacted with cardiolipin in a beta2-glycoprotein 1-dependent manner. The epitope for these antibodies consisted of beta2-glycoprotein 1 bound to cardiolipin or immobilized on plastic plates. The specificity is similar to the autoimmune anticardiolipin antibodies described in patients with SLE, APS and other autoimmune diseases. Monoclonal APA 510, 183, 238 reacted with cardiolipin in the absence of beta2-glicoprotein 1. beta2-Glicoprotein 1 , either in the fluid phase or bound to cardiolipin, inhibited the binding of these antibodies. Monoclonal APA 510 was cofactor-independent while monoclonal APA 183 and 238 reacted with cardiolipin only in the presence of human serum. The results of this study indicate that APA comprise a highly heterogeneous population of antibodies with respect to the antigens they recognize, as well as depending on presence of serum components.

[Cofactor-antiphospholipid antibodies relationship in patients with systemic lupus erythematosus and infertility women].

High frequency of detecting antiphospholipid antibodies (APA) is typical for patients with systemic autoimmune diseases as well as infertility women irresponsive to IVF treatment. Antiphospholipid antibodies cause thrombotic complications in patients with systemic autoimmune diseases contrary to antiphospholipid antibodies with in-vitro fertilization taken place. The authors have compared specificity and cofactor-antiphospholipid antibodies relationship in patient with systemic lupus erythematosus, infertility women irresponsive to IVF treatment and patients with chronic virus hepatitis. It has been shown that 80% of patients with systemic lupus erythematosus are mostly beta2-glycoprotein- and cofactor dependant. Patients with chronic virus hepatitis and women with in-vitro fertilization (IVF) have been found to have pretty low beta2-glycoprotein dependant antiphospholipid antibodies. Antiphospholipid antibodies in in-vitro fertilization women were cofactor dependant only in 28%. of the cases. These data can explain absence of thrombotic complication in APA positive IVF patient, because its complications have been associated with cofactor-dependent APA. It also can explain a lot of controversial results about significance of APA on IVF result.

[Investigations of fibrin clots formation under the effect of thrombin and ancystron]. / Doslidzhennia formuvannia fibrynovykh zgustkiv za diï trombinu ta antsystronu.

It was shown that heterogeneous surfaces of two phases influence on fibrin films structures. The clot formed between two phases have the surface layer which is different from fibrin clot surface structure. Transmission electron microscopy revealed to determine the thickness of the surface layer for desAA- and desAABB-fibrin. The surface layer thickness is 25,8 and 29 nm for desAA- and desAABB-fibrin respectively.

[Effect of progesterone and 17beta-estradiol on proinflammatory cytokine costimulatory proliferative activity]. / Vplyv prohesteronu ta 17beta-estradiolu na efekt kostymuliatsiï proliferatsiï prozapal'nymy tsytokinamy.

The lymphocyte proliferation is multicomponent mechanism of immune system reactivity. Many costimulatory factors take part in this process. Proinflammatory cytokines (TNF, IL-1 alpha and beta) enhance proliferation of activated lymphocytes. Female steroid hormones inhibit proliferation of mitogen and alloantigen-activated lymphocytes. The aim of this study was to investigate the effect of progesterone and 17 beta-estradiol on the costimulatory proliferative activity of proinflammatory cytokines in vitro. Female steroid hormones inhibit lymphocyte response to antiCD3 antibody. Progesterone had a stronger effect than 17 beta-estradiol (64 and 13% of inhibition respectively). 17 beta-estradiol enhanced the TNF costimulatory effect on the lymphocyte proliferation. Progesterone neutralized this TNF-induced effect and reverted it (inhibition of lymphocyte proliferation was enhanced in the presence of TNF). We found dominant inhibitory effect of progesterone on the TNF costimulatory activity when progesterone and estrogen were added simultaneously. Progesterone and 17 beta-estradiol downregulated costimulatory proliferative activity of IL-1 alpha or beta. Thus female steroid hormones had suppressive effect on the antiCD3-stimulated lymphocyte proliferation. They downregulated costimulatory proliferative activity of IL-1 alpha/beta and had opposite effect on TNF costimulatory activity. Our results suggest possible roles female steroid hormones as regulators on activity of proinflammatory cytokines and their functions in lymphocyte proliferation.

[Effect of female steroid hormones on expression of adhesion molecules by peripheral blood leukocytes]. / Ekspreciia molekul klitynnoï adheziï leikotsytamy peryferychnoï krovi pid vplyvom zhinochykh steroiïdnykh hormoniv.

The specific adhesion of cells to other cells or to extracellular matrices is a basic component of cell migration and recognition, and it underlies a lot of biologic processes including embryogenesis, tissue repair, and both immune and inflammatory responses. The aim of this study was to investigate the influence of female steroid hormones on expression of the adhesion molecules on the leukocytes. The whole blood from healthy people was incubated in a presence or an absence of progesterone (2 micrograms/ml) or 17 beta-estradiol (0.2 microgram/ml) for 4 h., and then with TNF for 18 h. The phenotype of the leukocytes was investigated by flow cytometry. Progesterone inhibited an expression of CD54 on monocytes and lymphocytes due to reducing density of these molecules on the cellular surface; 17 beta-estradiol inhibited an expression of CD54 on monocytes and CD69 molecules on monocytes and lymphocytes due to reducing density of these molecules on the cellular surface. Progesterone inhibited TNF-stimulated CD54 and CD11b expression on the granulocytes and CD69 expression on lymphocytes by reducing partly the density of these molecules on the surface of cells, and in such way it partly blocked the proinflammatory activity of this cytokine. Progesterone also reduced CD62L expression on the granulocytes by reducing an amount of a marker, positive to those cells but enhanced the effect of TNF. The data obtained evidence that female steroid hormones take part in the regulation of an expression of adhesion molecules by the leukocytes and are likely to be important in the circulation and activation of the leucocytes.

Immune disorders in women with premature ovarian failure in initial period.

PROBLEM: Premature ovarian failure (POF) may be considered as an autoimmune endocrine disease. Autoantibodies and lymphocyte subset changes are associated with premature ovarian failure. Immune cell parameters were studied in relation with anticardiolipin antibodies (ACAB) classes M and G in the initial period of POF. METHODS: Two-color flow cytometry was used to determine lymphocyte subsets and enzyme-linked immunosorbent assay (ELISA) was used to detect ACAB and hormones in the peripheral blood of 68 POF patients, 32 women with normal menopause (NM) and 13 healthy women as a normal control (NC). RESULTS: Patients in the initial period of POF had decreased levels of CD3+, CD19+, CD3+8+, and CD8+57+ lymphocytes and a high percentage of CD5 positive in CD19+ cell population compared to the control; frequencies of IgM ACAB in POF patients were significantly higher than both IgG ACAB and IgM ACAB in NC; correlation between lymphocyte subsets and hormone levels was absent. Women with early NM showed a low number of CD3+, CD3+4+, and CD3+8+ lymphocytes, a high number of CD3 + DR, and elevation of the percentage of CD5 positive in CD19+ lymphocytes compared with the control. The frequencies of both IgM and IgG ACAB were high; the levels of lymphocyte subsets had correlations with progesterone and estradiol concentrations. CONCLUSIONS: An increase of autoantibody producing B cells (CD5+19+) and a low number of effector suppressor/cytotoxic lymphocytes (CD8+57+) with active production of anticardiolipin autoantibodies class M were found. This suggested a primary autoimmune process in the initial period of POF. Autoimmune defeat of the ovary could be the primary cause of POF, whereas in NM autoimmunity is a result of hormone dysfunction.