Эпителиальные клетки тимуса и барьерных тканей секретируют цитокин, родственный интерлейки8 ну87 (IL87) - TSLP (Thymic stromal limphopoietin). Действуя на свою главную мишень - дендритные клетки, он индуцирует экспрессию мембранной молекулы OX40L. В процессе презентации антигена/ аллергена Т8лимфоциту через эту молекулу передается костимулирующий сигнал в CD4+ Т8клетку. Этот сигнал при условии отсутствия IL812 вызывает дифференцировку воспалительных Th28клеток, которые отличаются от обычных (регулирующих) Th28клеток тем, что наряду с типичными Th28ци8 токинами они вырабатывают фактор некроза опухоли, но не IL810. Результатом является индукция аллергического воспаления. Показана роль TSLP в развитии атопического дерматита и бронхиальной астмы. Формирование аллергических процессов можно предотвратить или ослабить, используя моно8 клональные антитела к OX40L, что предполагается использовать в качестве нового подхода к терапии аллергических заболеваний.

Cells with regulatory T-cell phenotype (Treg, CD4+CD25hi) were not detected in human fetal thymus, liver, bone marrow, spleen, and among blood mononuclears of 14–28-week gestation. The cells of the majority of these fetal thymuses express Treg specific marker (FOXP3 transcription marker) gene. Culturing of fetal liver and bone marrow cells on a monolayer of thymic epithelial cells induced expression of FOXP3 gene, but induction of CD4+CD25+ membrane phenotype was detected in only 1 of 8 studied cultures (in liver and bone marrow cells). Induction of Treg differentiation is to a greater extent determined by the characteristics of hemopoietic organ cells than of thymic epithelial cells.

Vilon stimulated and Epithalon suppressed the expression of argyrophilic proteins in nucleolar organizer regions of thymocytes and epithelial cells, stimulating or reducing, respectively, the formation, assembly, and transport of ribosomes into the cytoplasm and thus determining the intensity of protein synthesis in these cells. A direct mitogenic effect of Vilon was also revealed: this peptide promoted thymocyte transformation into proliferating blast cells.

OBJECTIVES The immunocytochemical study of the localization of hormones in thymic cells has been performed to clarify possible correlations of their expression with proliferative activity of thymocytes. METHODS We used commercial antibodies to serotonin, melatonin, somatostatin, glucagon, gastrin, beta-endorphin and histamine, and ABP or BSP kits for visualization of reaction. Computer image analysis was used to find correlations between hormone production and proliferative activity of thymocytes. RESULTS Different subpopulations of thymocytes are able to produce hormones: precursors of T-lymphocytes (CD4-CD8-) contain serotonin and melatonin, immature cortical cells (CD4+CD8+) produce only serotonin, mature medullar cells (CD4+CD8-) show immunoreactivity to serotonin, melatonin, beta-endorphin and histamine. The expression of serotonin, somatostatin and gastrin is localized in thymic epithelial cells, formatting Gassal's bodies. Proliferative activity of thymocytes depends from the expression of serotonin and somatostatin in thymic cells. CONCLUSION The data received testify the expression of different hormones in human thymic cells and showing by this fact high endocrine activity of thymus. The presence of correlations between hormonal expression and cell proliferative activity could be considered as the bright illustration of important role of neuroimmunoendocrine mechanisms in the regulation of local thymic homeostasis.

We studied the effects of the main placental hormone, chorionic gonadotropin, on differentiation of human thymocytes in vitro in the presence of thymic epithelial cells. It was shown that the hormone at a high dose (100 IU/ml) enhanced the epithelium-induced phenotypic maturation of thymocytes, which is registered by an increased expression of the membrane marker CD3 and transition of CD4 + 8 + thymocytes in the cells with CD4 + 8 – and CD4 – 8 + phenotypes. In addition, gonadotropin enhanced the proliferative response of thymocytes to the mitogen during their cultivation with the epithelium. The stimulating effect of the hormone on the epithelium-induced differentiation of thymocytes is mediated by the humoral factors of epithelial cells. In addition, gonadotropin at this dose exerts its own differentiating activity with respect to thymocytes and stimulates their phenotypic and functional maturation in a monoculture.

The main goal of this investigation was to evaluate the abnormal T-cell immunity in cleanup workers who took part in the cleanup after the Chernobyl accident in 1986. Peripheral blood mononuclear cells (MNCs) of apparently healthy cleanup workers (n = 134) were used to analyze the phenotype and proliferative response to mitogens in vitro. Evaluation of the MNC phenotype of cleanup workers did not reveal a significant disturbance in the T-cell subpopulation content except for an increase in CD3+CD16+56+ (NKT) cells. Immunophenotyping of phytohemagglutinin (PHA)-activated MNCs demonstrated suppression of CD4+ T-cell propagation and augmentation of CD8+ T-cell propagation in vitro compared to control individuals. DNA synthesis in the MNCs of cleanup workers was markedly inhibited after activation for 3 days with suboptimal concentrations of PHA, pokeweed mitogen and PMA. In contrast to control individuals, the monocytes of cleanup workers were able to stimulate the proliferation of T cells from healthy individuals but inhibited the proliferation of T cells from cleanup workers. This study affords a better understanding of the response of MNCs to stimulation with suboptimal concentrations of PHA and provides an approach to a more accurate analysis of the immunological disorders found after exposure to radiation from Chernobyl-related activities.

The ability of main reproductive hormones such as chorionic gonadotropin (CG), estradiol, and progesterone to regulate apoptosis of human neutrophils was studied. The hormones were studied separately and in physiological combinations specific for different trimesters of pregnancy. A low dose of CG (10 IU/ml) increased the spontaneous apoptosis of neutrophils, whereas its combination with estradiol and progesterone corresponding to that of trimester III of pregnancy significantly decreased this parameter. The stimulating effect of CG was prevented by an inhibitor of protein kinase A, whereas the hormone-induced suppression of apoptosis depended on the activity of Ca2+-channels. The antiapoptotic effect of the hormonal combination corresponding to that of trimester III was also manifested in the presence of autologous T-lymphocytes and on stimulation of neutrophils by bacterial lipopolysaccharide. The apoptosis induced with monoclonal antibodies to CD95 was significantly suppressed by the hormones studied and their combinations. Thus, apoptosis of neutrophils is effectively regulated by reproductive hormones; this seems to be an important control mechanism of activation of these cells in pregnancy.

Effects of chorionic gonadotropin (CG), estradiol, progesterone, and their physiological combinations on apoptosis of human peripheral blood T-lymphocytes were studied. Neither the hormones separately nor their combinations affected the spontaneous apoptosis of T-cells. On stimulation with mitogens, a high dose of CG (100 IU/ml) significantly increased apoptosis of T-lymphocytes, but its combination with steroid hormones specific for trimester I of pregnancy decreased this parameter. Apoptosis of T-lymphocytes induced by neutrophils in mixed culture was also inhibited by the hormone combination corresponding to trimester I. In greater detail, this hormonal combination was shown to display differential effects on different T-cell subpopulations: it stimulated apoptosis of CD8(+)-lymphocytes (which seemed to be provided by CG) and inhibited apoptosis of CD4(+)-cells. Apoptosis of T-lymphocytes induced by anti-CD95 was suppressed by a high dose of progesterone (100 ng/ml) and also by its combination with CG and estradiol specific for trimester III of pregnancy. Thus, the reproductive hormones studied effectively regulated apoptosis of peripheral blood T-lymphocytes. The effect of the hormones depended on the cell type and their activation and seemed to be an important mechanism of hormonal control of immune reactions in pregnancy.

The topography of thymocytes expressing neuronal and inducible nitric oxide synthases and changes in the content of luminescent immunoreactive products in these cells after intraperitoneal injection of bacterial lipopolysaccharide were studied by double immunohistochemical labeling. Under normal conditions neuronal nitric oxide synthase-immunopositive cells formed a wide network in thymus medulla (except for perivascular regions). Inducible nitric oxide synthase was expressed in single cells at the corticomedullary boundary. Lipopolysaccharide markedly increased the intensity of luminescence and number of inducible nitric oxide synthase-immunoreactive cells. However, this agent sharply decreased the intensity of luminescence in neuronal nitric oxide synthase-immunopositive cells of the stroma. Our results indicate that neuronal and inducible nitric oxide synthases are synthesized in various stromal cells of the thymus. Expression of these enzyme isoforms undergoes opposite changes during inflammation.

Co-cultivation of human thymocytes with homologous thymic epithelial cells (TEC) resulted in apoptosis of thymocytes and increase of CD25 expression. TEC supernatant also induced these effects. Fraction of apoptotic cells was enriched by CD69+ cells but not by CD95+ cells. Thymocytes of mice MRL-lpr/lpr bearing mutant form of gene Fas were sensitive to apoptosis induction in co-culture with TEC in the same degree as thymocytes of mice bearing Fas gene of wild type. Apoptosis of murine thymocytes can be induced by co-cultivation with both murine and human TEC.

AIM:To investigate the effect of recombinant alpha 2b-interferon (r alpha 2b-IFN) on functional capacity of peripheral blood (PB) T cells in rheumatoid arthritis (RA) patients and the relationship between functional characteristics of T lymphocytes and the disease activity.MATERIALS AND METHODS:PB mononuclear cells (PBMC) were separated by Ficoll-Verografine++ gradient centrifugation from 24 healthy donors (HD) and 75 RA patients 19 of which were treated with r alpha 2b-IFN (realdiron, Biofa, Lithuania) in the dosage 1 million IU i.m. each other day for 20 days, 10 injections a course. Cell surface markers (CD3, CD4, CD8) and adhesion molecules (CD18, CD54, CD2) were analyzed using specific monoclonal antibodies (MoAbs) and flow cytometry on the PBMC, freshly isolated and treated for 72 hours with medium alone, PHA, r alpha 2b-IFN and their combination. The proliferative response of PBMC to MoAbs for CD3, PHA and r alpha 2b-IFN were assessed by 3H-thymidine incorporation. The percentage of spontaneous and inducing apoptosis was quantified by flow cytometry using propidium iodide staining.RESULTS:The expression of CD18 was lower on RA PB lymphocytes compared to HD PB lymphocytes (p < 0.05). After stimulation of PBMC in both RA patients and HD with PHA, percentages of CD2+, CD3+, CD4+, CD18+ cells significantly diminished (p < 0.05), whereas the percentages of CD54+ and CD18+ (p < 0.05) cells increased. We have found three types of RA PB lymphocytes response to complex factors in vitro: 1) the presence of the proliferative response to T-mitogens but not to r alpha 2b-IFN (56% of the patients); 2) the presence of the increased proliferative response to T-mitogens and r alpha 2b-IFN (17% of the patients); 3) the absence of the proliferative response to T-mitogens and r alpha 2b-IFN (27% of the patients). PBMC of HD demonstrate only the first type of the response. R2 alpha b-IFN demonstrated own mitogenic effect and increased mitogen-induced proliferation in PBMC cultures with a high proliferative response to T-mitogens. The levels of spontaneous and inducing apoptosis were increased in RA PB lymphocytes compared to HD. After stimulation with PHA, RA PB lymphocytes preferentially underwent apoptosis whereas cells of HD proliferated. High disease activity correlated positively with an increase of a proliferative response to mitogens and apoptosis and a decrease in the percentage of lymphocytes, expressed adhesion molecules. The treatment with r alpha 2b-IFN induces changes in T-cell response to mitogens similarly to those after incubation with r alpha 2b-IFN in vitro before treatment.CONCLUSION:Functional capacity of RA PB lymphocytes relates to the disease activity. Inhibitory or stimulatory effects of r alpha 2b-IFN depend on functional activity of RA lymphocytes. Using the test with alpha 2b-IFN incubation, we may predict changes of apoptosis and proliferation levels caused by different agents in RA lymphocytes after treatment with r alpha 2b-IFN.

OBJECTIVES: Taking into account the hypothesis that Alzheimer's disease (AD) might be a systemic disease that affects several tissues in the body, the aim of this study was to try to detect the expression of tau-protein in human peripheral blood lymphocytes (PBL) in patients with AD. MATERIAL AND METHODS: Blood samples were obtained from patients with AD (n=16, age 67-98) and from volunteers without psychoneurological pathology (n=10, age 65-78). PBL were isolated on Ficoll-Paque gradient centrifugation. For cell fixation and permeabilization we used a fixative solution (4% formaldehyde and 0.1% glutaraldehyde) and 0.03% Triton X-100. Immunocytochemical detection of tau-protein was carried out by biotin-streptavidin complex method with tau monoclonal antibody (1:100, clone TAU-2, ICN) and universal immunostaining kit IMMU-MARK (ICN). RESULTS: The expression of tau-protein was shown in PBL in absolute majority of AD patients studied. Only in two healthy volunteers a single lymphocyte from many cells (i.e. a smear) demonstrated a very weak-positive immunostaining to tau-protein CONCLUSION: This first demonstration of clear difference in localization of tau-protein in blood lymphocytes between healthy and sick people testifies to the fact that tau-protein could be considered as a promising marker and blood lymphocytes as a suitable sample for life-time diagnosis of AD.

AIM:To elucidate the effects of combined therapy with glucocorticosteroids (GCS) and recombinant human interferon alpha or gamma (IFN) on proliferative responses of T-lymphocytes activated by various surface molecules (CD3 and CD2) in patients with SLE.MATERIALS AND METHODS:A 3-month trial entered 3 groups 15 patients each with verified SLE by APA criteria (1982). Patients of group 1, 2 and 3 received IFN-alpha (realdiron, Biofa, Lithuania) in a single dose 3 million IU i.m., IFN-gamma (inflagen, Biofa, Lithuania) in a single dose 3 million IU i.m. and cyclophosphamide in a dose 200 mg i.m. once a week, respectively. T-lymphocyte proliferative response was assessed to stimuli of two types: CD3-dependent (classic activation) and CD2-dependent (alternative activation). The analysis was made by inclusion of 3H-thymidine after 72-hour incubation of peripheral blood mononuclear cells with various stimuli. The response was assessed before the treatment, on the treatment day 20 and after the treatment. The blood from 27 donors was also examined. Flow cytometry estimated the percentage of the cells expressing molecules CD3, CD4, CD8.RESULTS:The effect is found of alpha and gamma INF on functional capacity of T-lymphocytes and on the number of cells expressing surface molecules CD3 and CD4. Realdiron produced two-phase reaction to a proliferative response to mitogenic stimuli by CD3-dependent activation pathway: the initial rise then lowering. CD2-dependent way of T-cell activation is associated with weakening of responses to all combinations of stimuli with participation of autologous red cells. This group of patients to the end of the therapy exhibited a significant decrease in the number of cells expressing CD3 and CD4 (p < 0.05 and p < 0.001, respectively). Inflagen enhanced CD3-dependent activation of T cells and normalized the response to all types of the alternative stimuli. This group demonstrated an increase in the number of cells expressing CD3 and CD4 (p < 0.01 and p < 0.05, respectively). The changes in the number of CD8+ cells in both the groups were statistically insignificant. The controls had T-cell responses reduced by both activation pathways.CONCLUSION:Preparations of both alpha and gamma interferon have a multidirectional influence on functional potential and phenotype of T-lymphocytes of SLE patients.

AIM:To study activation of T-lymphocytes by the CD3 (antigen-dependent) and CD2 (non-antigen-dependent) routes in patients with systemic lupus erythematosus (SLE).MATERIALS AND METHODS:Peripheral blood mononuclears were studied in 66 patients with SLE and 27 donors. Proliferative response to activation by anti-CD3, anti-CD3+ phorbol-12-myristate-13-acetate (PMA), phytohemagglutinin (PHA), and autologous erythrocytes in combinations with PMA and recombinant interleukin-2 (rIL-2) was assessed.RESULTS:T-cell proliferation was at least two times increased under the effect of CD3 in 40.9% patients and in 100% normal subjects. Stimulation with CD3 antibodies in combination with PMA leveled the differences due to boosting of T-cell response in SLE patients. PMA alone caused mononuclear proliferation in 25% patients with SLE but not in normal subjects. Decreased response of T-cells to adhesive stimulus (autologous erythrocytes + PMA) in SLE patients was leveled by rIL-2.CONCLUSION:The proliferative response of T-lymphocytes is decreased upon stimulation with CD3 and CD2 and in some patients increased by PMA in submitogenic doses, added alone or in combination with anti-CD3.

Apoptosis, together with proliferation, is a main factor of selection of the clones of developing T-lymphocytes: the clones not supported by positive selection are subject to apoptosis and apoptosis accounts for discarding of potentially autoaggressive clones, i.e., for negative selection in the thymus and peripheral lymphoid tissue. Realization of apoptosis at different stages of the development of T-lymphocytes depends to a varying extent on Fas, Bcl-2, p53, and other regulators. The dendritic cells are the main cell type, the contact with determines apoptosis of T-lymphocytes. A possible role of the epithelial cells was shown in few models (on murine cells) and was not practically studied. We obtained a line of epithelial cells of the human thymus cells HTSC, cocultivation with which induces apoptosis of immature thymocytes and blood T-cells activated by mitogens. Development of apoptosis is suppressed by inhibitors of protein and RNA synthesis, chelators Ca2+, ions Zn2+ and factors destroying the cytoskeleton components. In this model, interaction of pairs of molecules CD4-HLA class II and LFA-1-ICAM-1. When in contact with the HTSC cells, the thymocytes of mice mutant for Fas-receptor (line MRL.lpr) are subject to apoptosis, but when this receptor is present, it affects the development of apoptosis.