Spontaneous Cytokine Production Research Articles

Spontaneous and Stimulated Production of Cytokines by Blood Cells Ex Vivo as a Biomarker of Initially High or Low Hypoxia Resistance in Rats.

Spontaneous and stimulated production of cytokines by peripheral blood cells obtained from the caudal vein of male Wistar rats was assessed before testing their resistance to oxygen deficiency in a decompression chamber. To study the spontaneous production of cytokines, heparinized blood cells were incubated in a culture medium (24 h, 5% CO2, 37°C) and the content of proinflammatory cytokines IL-6 and TNFα and anti-inflammatory IL-10 in the culture medium was assessed by ELISA. To stimulate cytokine production, blood cells were incubated for 24 h with LPS, phytohemagglutinin, and concanavalin A at final concentrations of 2, 4, and 4 μg, respectively. Two weeks after blood sampling, individual resistance of the animals to hypoxia in a decompression chamber was determined. In animals with low resistance to hypoxia, the levels of spontaneous production of all three cytokines were significantly higher, while after stimulation, the level of IL-1β increased by more than 2 times. The animals with spontaneous production of IL-10>50 pg/ml, IL-6>10 pg/ml, and TNFα>10 pg/ml, as well as with the increase in IL-1β production by more than 2 times upon stimulation were classified as low-resistant. At IL-10<15 pg/ml, IL-6<9 pg/ml, and TNFα<7 pg/ml, as well as the absence of the increase in IL-1β production upon stimulation, they were classified as high-resistant. The identified features of spontaneous and stimulated production of cytokines can be used as non-invasive biomarkers to determine the resistance to hypoxia without exposure to sublethal hypoxia in a decompression chamber.

Spontaneous cytokine production by the cells from peripheral blood, lymph nodes, spleen, thymus and skin in a murine model of acute psoriasis-like dermatitis

Imiquimod-induced model of psoriasis in mice is used for the studies in psoriasis. Imiquimod administered as skin application triggers a complex multi-stage process simulating psoriatic inflammation of the skin, accompanied by changes in production of cytokines. The aim of this work was a comprehensive study of their contents in cell supernatants isolated from skin, blood, from central and peripheral lymphoid organs and the effect of imiquimod on these parameters. Forty-six C57BL/6 mice were divided into two groups: (1) control (n = 22), and (2) experimental (n = 24). Development of experimental pathology was evaluated by van de Fits et al. The mice from experimental group were treated with a cream containing 5% imiquimod (62.5 mg/ cm2/ day/mouse) for 7 days. On the 7th day of experiment, the animals were subject to blood sampling, extraction of spleen, lymph nodes, thymus, and skin biopsies were also made. To obtain cell suspensions, the tissues of spleen, lymph nodes, and thymus were crushed in a homogenizer, and then passed through cell filters (50 mcm pore size). The method of spontaneous migration was used to isolate skin cells. The cultures of blood, spleen, lymph nodes, thymus, and skin cells were incubated for 24 hours in RPMI-1640 followed by assessment of spontaneous cytokine production in supernatants. The cytokine level (IFNγ, IL-1β, IL-5, IL-6, IL-10, IL- 17, TNFα, IL-10) was determined using the murine Th1/Th2/Th17 Panel test system. Our study revealed that the cells isolated from blood and lymphoid organs synthesize increased amounts of pro-inflammatory cytokines: IL-1 and IL-17. The most pronounced changes in the cytokine profile are observed in cell supernatants from blood, skin and spleen cultures.

Assessment of the effect of using immunomodulatory drugs for emergency prevention of experimental plague caused by a virulent strain of the main subspecies Yersinia pestis

Introduction. Immunomodulatory drugs (IMD) have great potential to increase the nonspecific reactivity of the organism in a set of measures for emergency prevention of plague. The purpose of the work is to evaluate the protective effectiveness of the use of IMD of different groups in experiments on modeling infection with a highly virulent strain of the plague microbe. Materials and methods. IMD (rIFN-γ — recombinant interferon-gamma, PO — azoximer bromide, synthetic immunomodulatory oligopeptides O1, O2, O3) was administered to white mice and guinea pigs subcutaneously thrice by the schedule 3 days, 1 day, and 1 hour prior the infection with a virulent test strain of plague Yersinia pestis 231 (708) at dose from 1 to 625 CFU. In addition, the effect of IMD on the production of IFN-γ and interleukin-10 was investigated in white mice before infection. Results. The study of the effect of IMD on the survival of unvaccinated biomodels made it possible to establish that only rIFN-γ and PO increase the survival of two types of laboratory animals by 20–50% and significantly increase the LD50. All tested IMD contribute to an increase in the average life expectancy of biomodels by at least one day. An increase in spontaneous and mitogen-induced cytokine production was found only in white mice receiving rIFN-γ and PO, which correlates with animal survival rates. Conclusion. The obtained data indicate the effectiveness of the use of IMD, especially rIFN-γ and PO in protecting the macroorganisms from infection with Y. pestis, which determines the prospects for research on the further improvement of emergency prevention of plague.

Spontaneous and LPS-induced cytokine production in supernatants of cell cultures isolated from a skin biopsy in a mouse model of acute psoriasis-like dermatitis

The purpose of the work performed was to obtain data on spontaneous and LPS-stimulated cytokine production in cell culture supernatants isolated from isolated from skin bioptates of C57BL/6 mice in an imiquimod-induced psoriasis model. Methods. Skin cells were isolated by spontaneous migration and enzymatic dissociation in a collagenase solution. Cells isolated from the skin were cultured for 24 hours in RPMI-1640 (spontaneous cytokine production) or RPMI-1640 medium with the addition of a solution (10 micrograms/ml) of E. coli LPS (induced). Cytokine levels were determined by flow laser cytofluorometry using the Mouse Th1/Th2/Th17 multiplex test system (Antigenix America, USA) on an FC-500 cytometer. Results. In the focus of imiquimod-induced inflammation, an increase in the spontaneous production of proinflammatory cytokines IL-1, IL-5, IL-17 was observed by 2.4-3.8 times. At the same time, a 1.9-fold inhibition of IFNy synthesis was detected when cultivating skin cells in the presence of bacterial LPS.

The ecto-enzyme CD38 modulates CD4T cell immunometabolic responses and participates in HIV pathogenesis.

Despite abundant evidence correlating T cell CD38 expression and HIV infection pathogenesis, its role as a CD4T cell immunometabolic regulator remains unclear. We find that CD38's extracellular glycohydrolase activity restricts metabolic reprogramming after T cell receptor (TCR)-engaging stimulation in Jurkat T CD4 cells, together with functional responses, while reducing intracellular nicotinamide adenine dinucleotide and nicotinamide mononucleotide concentrations. Selective elimination of CD38's ectoenzyme function licenses them to decrease the oxygen consumption rate/extracellular acidification rate ratio upon TCR signaling and to increase cycling, proliferation, survival, and CD40L induction. Pharmacological inhibition of ecto-CD38 catalytic activity in TM cells from chronic HIV-infected patients rescued TCR-triggered responses, including differentiation and effector functions, while reverting abnormally increased basal glycolysis, cycling, and spontaneous proinflammatory cytokine production. Additionally, ecto-CD38 blockage normalized basal and TCR-induced mitochondrial morphofunctionality, while increasing respiratory capacity in cells from HIV+ patients and healthy individuals. Ectoenzyme CD38's immunometabolic restriction of TCR-involving stimulation is relevant to CD4T cell biology and to the deleterious effects of CD38 overexpression in HIV disease.

Systemic production of cytokines and growth factors in rats with metabolic syndrome

ABSTRACT. An important characteristic of the state of the immune system is the production of cytokines, which determine the development of metabolic processes and are aimed at the immune response. Changes in the immune response occur with the development of homeostasis system disorders that change the rheological characteristics of the blood and contribute to the development of the metabolic syndrome (MS). The aim of the study was to study changes in the functional activity of cytokines and angiogenic growth factors (GF) during experimental MS in rats depending on age. Мaterials. MS modeling was carried out on 21 rats, which were divided into 3 groups, 7 animals each, of the same age (1a subgroup - 6 months of age, 2a subgroup - 9 months of age, 3a subgroup - 12 months). In the pathogenesis of the development of MS The role of MS in the development of systemic production of cytokines and FR has been studied, and data on the content of pro-inflammatory cytokines in the peripheral blood depending on the age of rats have been presented. The results of the study of the content of growth factor (GF) and endothelin in peripheral blood in MS tended to decrease with increasing age of rats providing morpho- and angiogenesis (p&lt;0.01). An increase in the concentration of endothelin in the blood serum of rats with MS may contribute to the activation of vascular endothelial cells and an increase in the level of fibrinoid in the vascular system with increasing age. The level of cytokines circulating in the peripheral blood in rats with MS significantly differed from that, which was manifested by an increase in the level of IL-4, IL-8 and IL-10. Spontaneous production of cytokines by peripheral blood lymphocytes in MS increased with the age of rats, which indicated an increase in the secretion of IL-6 and IL-10, and a decrease in the production of IFN-1 and TNF-α. The decrease in spontaneous production of IL-1β and IL-10 in 24-hour cultures of peripheral blood monocytes in rats was significantly higher compared to IL-6 (p&lt;0.001) compared to the 2nd and 3rd - observation groups. The content of IL-8 and TNF-α did not differ significantly (p&gt;0.05), and IL-12 was below the sensitivity limit of the test system (less than 3 pg/ml). Conclusion. The data obtained indicate that in the peripheral blood of rats with MS, the secretion of pro-inflammatory and regulatory cytokines, as well as FR, decreases with age. Hormones also have an inhibitory effect on the production of cytokines by macrophages, the level of which changes significantly during MS. The results of the study indicate various pathogenetic mechanisms of the formation of MS in rats, accompanied by a violation of the immune response of mononuclear cells and angiogenic RF at the system level.

Human endogenous retrovirus HERV-E λ 4-1 in immunopathogenesis of affective disorder

The aim of this work was to study a dependence between the production level of some pro-inflammatory cytokines by peripheral blood mononuclear cells, and activation of human endogenous retrovirus HERV-E 4-1 in the patients with recurrent depression. Patients and methods: the study included 30 patients with an verified diagnosis of recurrent depression (F 33.0) aged 26-45 years, with a disease duration of at least 3 months prior to inclusion into the study. Peripheral blood mononuclear cells were isolated by centrifugation in Ficoll density gradient (1.078 g/cm3). The human endogenous retrovirus HERV-E 4-1 env gene expression was determined by polymerase chain reaction using paired oligonucleotide primers. To assess the cytokine production, peripheral blood mononuclear cells were cultured for 24-72 hours, depending on the experimental conditions. Quantitative determination of spontaneous cytokine production was carried out by a sandwich variant of ELISA method in conditioned media from the cell cultures, according to the manufacturer instructions. Results: our data reveal higher production of IL-1 and IFN in peripheral blood mononuclear cells from those patients with recurrent depression who showed detectable HERV-E 4-1 env expression compared to the patients in whom the HERV-E 4-1 env gene expression was not detected. When studying correlation between HERV-E 4-1 env expression and production of IL-1 and IFN, a positive correlation between the studied parameters was established. Thus, taking into account our earlier data on HERV-E 4-1 immunomodulatory properties, as well as literature data concerning the HERV transcripts found in brains of mentally ill patients, along with increase of IL-1 and IFN production in the patients with recurrent depression and positive HERV-E 4-1 env gene expression, and a positive correlation between the HERV-E 4-1 env gene expression and increased level of cytokines involved in formation of pathological events in the nervous system in the patients with depression, one may conclude that activation of HERV-E 4-1 could participate in immunopathogenesis of recurrent depression by stimulating the synthesis of pro-inflammatory cytokines.

Concentration of anti-inflammatory cytokines in supernatants of peripheral blood cell cultures in children with autoimmune and infectious diseases

Cytokines belong to the class of signaling molecules, being involved into regulation of proliferation, differentiation and effector functions of immunocompetent cells. The ratio of pro- and anti-inflammatory cytokines is important for development of any inflammatory process. IL-1ra, IL-4, IL-10 are among the most important anti-inflammatory cytokines. Their function is to limit and suppress immune response in inflammatory processes of any etiology. In this respect, the aim of this study was to evaluate production of the anti-inflammatory cytokines IL-1ra, IL-4 and IL-10 by peripheral blood cells in children with autoimmune and infectious diseases. Patients and methods: Pediatric parients (2-17 years old) participated in the study including those with juvenile idiopathic arthritis (n = 101); unspecified reactive arthropathy (n = 24); systemic lupus erythematosus (SLE, n = 14); chronic viral hepatitis C (n = 24). 33 healthy children (n = 33) comprised the control group. Heparinized blood samples were diluted with glutamine-containing medium RPMI-1640. Control samples were not treated by any stimulants, and the stimulated samples were supplied with phytohemagglutinin (20 mkg/ml). The samples of diluted blood were incubated for 24 hours (37 C, 5% CO2). The supernates were frozen once. The concentrations of IL-1ra, IL-4 and IL-10 in these cell supernatants were determined by ELISA technique (Vector-Best, Russia). Results: It was found that the spontaneous production of anti-inflammatory cytokines (IL-1ra, IL-4 and IL-10) didnt differ, or was lower in the groups of patients with SLE, juvenile idiopathic arthritis and hepatitis C if compared with control group. SLE is an autoimmune disease, whereas juvenile arthritis is of mixed autoimmune-autoinflammatory etiology. Chronic hepatitis C is a viral disease, but autoimmune responses may manifest at the chronic stage of the disorder. Decreased production of anti-inflammatory cytokines could be an evidence for autoimmune mechanisms of these diseases, despite their different etiology. More intensive spontaneous production of IL-1ra and IL-10 in children with unspecified reactive arthropathy may suggest some compensatory reactions which inhibit development of inflammation in this disorder. IL-1ra, IL-4 and IL-10 production in stimulated cultures didnt differ between all groups of the patients, or it was lower in comparison with healthy children. Decrease cytokine production in groups of children with different diseases suggests exhausted functional reserve of immunocompetent cells caused by their chronic activation.

Cytokine Production in Whole Blood Cells Culture of Patients with Alcohol Dependence and Autologous Plasma Oxidative Stress Markers

We studied spontaneous production of a spectrum of proinflammatory cytokines by cultured whole blood cells from men with alcohol dependence at the stage of withdrawal syndrome and oxidative stress markers (carbonylated proteins and TBA-reactive substances) in the plasma of these blood samples. Enhanced production of cytokines by blood cells and increased concentrations of oxidative stress markers in the autologous plasma were revealed in comparison with the corresponding parameters in the control (blood from healthy men). Direct correlations were found between the levels of spontaneous cytokine production by blood cells from subjects with alcohol dependence and the concentration of oxidized proteins and lipids in autologous plasma.

Evaluation of cytokine levels in response to mitogen among HIV-1-infected blood cells and their relationships to the number of T cells

BackgroundHuman immunodeficiency virus-1 (HIV-1) infection causes loss and anergy of CD4+ and CD8+ T cells, leading to opportunistic infections, including tuberculosis (TB). QuantiFERON®-TB (QFT) is used as a diagnostic tool to detect TB, but it exhibits limited accuracy among subjects with low CD4+ T cell numbers, including HIV-1-infected individuals. The present study aimed to determine the effect of HIV-1 infection and patients’ blood T cell numbers on cytokine production in response to mitogen (Mit) stimulation. MethodsThe number of CD4+ and CD8+ T cells in HIV-1-infected individuals was quantified. Levels of various cytokines in Mit-stimulated and un-stimulated (Nil) supernatants of QFT gold “in tube” were assessed using a MAGPIX System. The correlation between cytokine levels and CD4+/CD8+ T cell counts in response to Mit was analyzed. The cytokine levels were compared between HIV-1-infected and healthy subjects. ResultsHIV-1-infected individuals (110) and control subjects (27) were enrolled. Interferon (IFN)-γ, interleukin-1 receptor antagonist (IL-1RA), IL-6, IL-8, and regulated on activation, normal T cell expressed and secreted (RANTES) values in Mit-Nil tubes showed a significant correlation with CD4+ T cell counts, while IFN-γ, IL-6, and IFN-γ-induced protein 10 (IP-10) values in Mit-Nil tubes had significant correlation with CD8+ T cell counts. IL-1RA, IL-8, IP-10, platelet-derived growth factor (PDGF)-BB, and RANTES levels in Nil tubes were significantly higher in the HIV-1-infected group. IFN-γ, IL-2, IL-5, IL-6, IP-10, and macrophage inflammatory protein-1β values in Mit-Nil tubes were significantly higher, and PDGF-BB and RANTES levels were significantly lower in the HIV-1-infected group. ConclusionThe functions of HIV-1-infected T cells and uninfected T cells, such as spontaneous and responsive cytokine production in response to Mit, were different. Our findings may be useful for developing new clinical tools for patients with low T cell counts. Additionally, the study provides new insights into the pathogenesis of HIV-1 infection.

РОЛЬ ИММУНОЛОГИЧЕСКИХ ФАКТОРОВ В НЕБЛАГОПРИЯТНОМ ИСХОДЕ ХРОНИЧЕСКОЙ ОБСТРУКТИВНОЙ БОЛЕЗНИ ЛЕГКИХ

Aim. The aim of the research was to study the role of immunological mechanisms in the unfavorable outcome of chronic obstructive pulmonary disease (COPD). Material and methods. The research included 116 patients hospitalized in 2005-2006 in the pulmonology department of patients with the exacerbation of mild and moderate COPD. Research protocol: general clinical and special (immunological) methods of examination were performed in patients with COPD. Instrumental research methods included: spirography, ECG, EchoCG, fibrobronchoscopy, chest X-ray. After diagnostic bronchoscopy, bronchoalveolar fluid (BAF) was taken for cytological and immunological examination. On the first and second days of inpatient treatment, patients underwent immunological studies: immunophenotyping of mononuclear cells (MNC), assessment of phagocytic activity of leukocytes (latex test, nitroblue tetrazolium (NBT) test) in blood and BAF, determination of the content of immunoglobulins in blood serum and BAF, study of cytokine levels in blood serum, BAF and MNC culture supernatant under conditions of their spontaneous production and activation in the mitogen-stimulated (phytohemagglutinin "Difco" 5 mg/ml) proliferation. Statistical data processing was carried out using the Statistica 10.0 program. In 2020, the long-term survival of COPD patients was assessed. The cohort of patients was divided into two groups. Survivors were included in the first group (n=44), and patients who died by 2020 were included in the other group (n=72). The retrospective comparison of the studied indicators was determined at the time of COPD exacerbation in 2005- 2006 in these patient groups. Results and discussion. In the surviving patients, the BAF cytogram was distinguished by the higher macrophage content and the increased number of NBT-positive cells in this population with the reduced neutrophil content in both blood and BAF. The deceased had elevated levels of IgM and IgA in the blood serum and BAF. The results of the study of cytokine production in MNC cultures in vitro indicate that the ratio of IFN-γ/IL-4 in the conditions of spontaneous cytokine production was higher in the deceased compared with the indicators of the surviving patients. This fact indicates the polarization of the immune response of the deceased in the direction of the cell type mediated by Th1 cells, which is confirmed by the peculiarities of the BAF cytokine profile in the deceased – the significant increase in the value of IFN-γ/IL-4 relative to the indicators in the survivors. Conclusion. The unfavorable outcome of COPD is associated with the increase in the number of neutrophil cells in BAF, in the blood, with the predominance of Th1-type activation of adaptive immunity at the local level and activation of both cellular and humoral mechanisms of adaptive immunity at the systemic level. The increased activity of innate immunity in COPD exacerbation, manifested by the increase in the number and metabolic activity of macrophages, is associated with the long-term survival of patients.

Ex vivo mass cytometry analysis reveals a profound myeloid proinflammatory signature in psoriatic arthritis synovial fluid

ObjectivesA number of immune populations have been implicated in psoriatic arthritis (PsA) pathogenesis. This study used mass cytometry (CyTOF) combined with transcriptomic analysis to generate a high-dimensional dataset of matched...

Experience of Using a Complex Antigenic Preparation of the Plague Microbe to Assess the Severity of a Specific Anti-Plague Response

Background. Improving the methodology of immunological monitoring in natural foci of plague in the Russian Federation and adjacent territories to increase the effectiveness of epidemiological surveillance of plague is an urgent line of research. The lack of correlation between the production of specific antibodies to the capsular antigen (F1) ofthe plague microbe with other indicators of the state of cellular defense reactivity indicates the need to search for new informative and accessible markers for assessing anti-plague immunity.Objective: to evaluate possibility of using the complex preparation (F1 and cell membranes) evaluate the possibilities of using an artificial antigenic complex based on F1 and cell membranes (CM) of the plague microbe in antigen-specific tests in vitro in people vaccinated against plague.resu. The study involved 153 volunteers living in the territory enzootic for plague (the village of Khandagayty ofthe Ovyur kozhuun of the Tyva Republic and the village of Kosh-Agach of the Kosh-Agach district of the Altai Republic). The study included the determination of spontaneous and mitogen-induced production of cytokines (IFN-γ, IL-4, TNF-α) by blood cells, titers of specific IgG antibodies to the capsular antigen F1 of the plague microbe and concentrations ofthe main classes of immunoglobulins (IgM, IgG, IgA and IgE) in blood serum, as well as immunophenotyping of blood lymphocytes (CD3, CD4, CD8, CD16, CD19).Results. Comparative assessment of the level of cytokines (TNF-α, IFN-γ and IL-4) in spontaneous/induced F1+CM Y. pestis tests revealed a statistically significant increase in the production of cytokines TNF-α and IFN-γ in the antigeninduced tests compared with spontaneous (p &lt; 0.01).Conclusion. Thus, the effectiveness of the use of artificial antigenic complex based on F1 and cell membranes ofthe plague microbe has been shown to assess the production of cytokines in antigen-specific cell tests in vitro, which justifies the need for further research.

Effects of long-term therapy with quetiapine and olanzapine on parameters of immunity and cytokine levels in patients with schizophrenia

IntroductionThe study of effects of long-term antipsychotic therapy in patients with schizophrenia is relevant.ObjectivesTo study effects of long-term antipsychotic therapy on parameters of immunity and cytokine levels in patients with schizophrenia.MethodsWe examined 20 schizophrenic patients, who received quetiapine (group 1) and 17 - olanzapine (group 2) for more than 6 months before admission in the hospital as the main anti-recurrence therapy. Persons aged 20-63 years with length of the follow-up of the disease ≥1 year were included. The investigations included: phenotyping of immunocompetent cells into CD differentiation clusters by flow cytometry; mitogen-induced, spontaneous production of cytokines (IL2, IFN-γ, IL-4, TNF-α) were identified with use of kits for enzyme-linked immunosorbent assay (ELISA).ResultsIt was shown that patients of group 1 in comparison with group 2 were characterized by lower values of СD3- lymphocytes (p=0.049), higher values of the spontaneous production of IFN-γ (p=0.01), mitogen-induced production of IL-2 (p=0.043) and IL-4 (p=0.059). In all examined low level of mitogen-induced of IFN-γ (p=0.0001) and TNFα (p=0.002; p=0.0001), high level of spontaneous production of TNFα (p=0.001) were revealed in relation to control.ConclusionsIt was found that the acute period of schizophrenia after prolonged treatment with atypical antipsychotics is accompanied by immunological imbalance and dysregulation of the cytokine system. More severe immune disorders when hospitalized during the exacerbation period were revealed in patients who had been receiving antipsychotic therapy with the atypical antipsychotic quetiapine for a long time. This can be associated with the features of the mechanism of action of atypical antipsychotics.DisclosureNo significant relationships.

Influence of immunomodulation on intracellular cytokine expression by spleen T-helpers of mice immunized by Yersinia pestis EV NIIEG

Aim. To characterize the intracellular expression of cytokines by spleen T-helpers and the spontaneous production of cytokines in the blood of BALB/c mice immunized with Yersinia pestis EV NIIEG against the background of immunomodulation.Materials and methods. Intracellular expression of CD4+IFN-γ+, CD4+IL-4+, CD4+IL-17+ was determined in mice spleen cell suspensions by flow cytometry, IFN-γ and IL-10 were measured in ELISA in blood supernatants on day 3 and day 21 after the immunization with Y. pestis EV against the background of immunomodulation. On day 21 after the immunization animals were infected by Y. pestis 231 at a dose of 400 LD50.Results. Differences in cytokine response to studied drugs, correlated with CD4+IFN-γ+ levels in animals, were identified. On day 3, a significant decrease in CD4+IFN-γ+ was observed in response to Y. pestis EV and to recombinant gamma interferon (Ingaron). A significant increase in CD4+IFN-γ+ was detected in response to vaccine strain administered with azoximer bromide (Polyoxidonium). Intracellular expression of IFN-γ, IL-4 and IL-17 increased on day 21by an average of 2,3 times when immunomodulators were used in the immunization schedule. In addition, on day 21 a significant (p ˂ 0.05) increase in the proportion of T-helpers expressing IFN-γ, as well as in level of spontaneous IFN-γ production in blood supernatants was observed only in animals immunized by schedules that included immunomodulators. After the challenge with Y. pestis 231 of animals previously immunized by schedules that included Polyoxidonium, the correlation analysis confirmed the association (r = 0,94; p = 0,0004) of mice survival with intensity of CD4+IFN-γ+ expression.Conclusion. The data obtained confirm the effectiveness of Polyoxidonium application in experimental animal Y. pestis EV immunization schedule and the usefulness of intracellular cytokine expression measurement for assessment of the level of protection following the immunization.

ROLE OF ADIPOSE TISSUE AND ADIPOKINES IN CHRONIC INFLAMMATION DEVELOPMENT ON METABOLIC SYNDROME BACKGROUND

The most pressing issue that combines obesity and insulin resistance is chronic subclinical inflammation, which affects the metabolic and secretory functions of adipose tissue, and is important for the development of pathological processes. The morphological basis of inflammation is the infiltration of adipose tissue by immune competent cells. Biologically active substances specific for adipose tissue are considered to be the collagen−like protein adiponectin and the protein hormone leptin, which are secreted in adipocytes. Leptin stimulates the cellular immune response and increases the production of pro−inflammatory cytokines, and adiponectin is thought to have anti−inflammatory properties. With the development of metabolic syndrome, the concentration of adiponectin in blood decreases, and that of leptin increases. To establish the relationship between serum leptin levels with markers of systemic inflammation and spontaneous production of proinflammatory cytokines as well as mononuclear blood leukocytes, an experimental study was conducted, i.e. modeling the metabolic syndrome in white female rats WAG / GSto aged 5−6 months. The predominance of proinflammatory cytokines: interleukins − 1β, −6, −8, −10, TNF−α in supernatants of mononuclear leukocytes with increasing leptin concentration, which is consistent with the view of its ability to stimulate cell immunity and affect the production of proinflammatory cytokines. It is proven that an increase in leptin levels in metabolic syndrome is not only a symptom that characterizes the functional state of adipose tissue, but also causes spontaneous production of proinflammatory cytokines and mononuclear leukocytes in blood, that is pathogenetically interrelated with the systemic inflammatory response. It is established that the change in the cytokine profile in the serum becomes a forecast of the formation and effectiveness of treatment of metabolic syndrome on the background of obesity. Key words: obesity, metabolic syndrome, undifferentiated chronic inflammation, cytokines.

Association of Maternal Factors and HIV Infection With Innate Cytokine Responses of Delivering Mothers and Newborns in Mozambique.

Maternal factors and exposure to pathogens have an impact on infant health. For instance, HIV exposed but uninfected infants have higher morbidity and mortality than HIV unexposed infants. Innate responses are the first line of defense and orchestrate the subsequent adaptive immune response and are especially relevant in newborns. To determine the association of maternal HIV infection with maternal and newborn innate immunity we analyzed the cytokine responses upon pattern recognition receptor (PRR) stimulations in the triad of maternal peripheral and placental blood as well as in cord blood in a cohort of mother-infant pairs from southern Mozambique. A total of 48 women (35 HIV-uninfected and 13 HIV-infected) were included. Women and infant innate responses positively correlated with each other. Age, gravidity and sex of the fetus had some associations with spontaneous production of cytokines in the maternal peripheral blood. HIV-infected women not receiving antiretroviral therapy (ART) before pregnancy showed decreased IL-8 and IL-6 PRR responses in peripheral blood compared to those HIV-uninfected, and PRR hyporesponsiveness for IL-8 was also found in the corresponding infant’s cord blood. HIV infection had a greater impact on placental blood responses, with significantly increased pro-inflammatory, TH1 and TH17 PRR responses in HIV-infected women not receiving ART before pregnancy compared to HIV-uninfected women. In conclusion, innate response of the mother and her newborn was altered by HIV infection in the women who did not receive ART before pregnancy. As these responses could be related to birth outcomes, targeted innate immune modulation could improve maternal and newborn health.

Transportation and Routine Veterinary Interventions Alter Immune Function in the Dog

Rapid activation of the hypothalamic pituitary adrenal axis and the sympathetic nervous system are hallmarks of the acute stress response and these systems interact with the immune system by signaling though glucocorticoid and adrenergic receptors on immune cells. There is limited information about the effect of these physiologic responses on immunologic parameters of pet dogs enrolled in clinical studies. The objective of this study was to evaluate how travel, instrumentation, and hospitalization alter immunologic parameters in pet dogs. Blood was collected from healthy dogs in a home environment and from healthy dogs at the time of presentation to the hospital and after instrumentation and 24 hours of hospitalization. We found that lipopolysaccharide (LPS)-induced downregulation of toll like receptor 4 (TLR4) was blunted in dogs exposed to stress. Neutrophil and monocyte major histocompatibility complex class II (MHCII) expression increased after transportation to the veterinary hospital but then became similar to that of the control dogs at the end of hospitalization. Peripheral blood mononuclear cell cytotoxicity function was blunted in dogs exposed to the stress of transportation as well as hospitalization. Neutrophil apoptosis was greater in dogs exposed to stress compared to controls although this effect significantly decreased after hospitalization stress. Conversely, stress did not alter induced or spontaneous cytokine production from leukocytes, neutrophil or monocyte expression of TLR4, LPS-induced downregulation of monocyte TLR4, LPS-induced neutrophil and monocyte expression of MHCII or peripheral blood lymphocyte phenotype. Transportation and instrumentation/hospitalization stress should be considered when interpreting immunologic studies in pet dogs.

P098 Receptor-interacting protein 1 kinase inhibition therapeutically ameliorates experimental T-cell-dependent colitis in mice

Abstract Background Inflammatory bowel disease (IBD) is a multifactorial disorder characterised by chronic and relapsing intestinal inflammation. Receptor-interacting protein 1 (RIP1) kinase activity is emerging as a driver of pro-inflammatory cytokine production and cell death and has been implicated in multiple inflammatory diseases including IBD. To this end, recent work has shown that RIP1 kinase inhibition reduces the spontaneous production of cytokines from human ulcerative colitis (UC) and Crohn’s disease explants. Methods The aim of this study was to investigate the anti-inflammatory effect of a highly selective inhibitor of RIP1 kinase activity (GSK547A) in the mouse T-cell transfer model of colitis; a model that shares many features with human IBD. All animal studies were ethically reviewed and carried out in accordance with Animals (Scientific Procedures) Act 1986 and the GSK Policy on the Care, Welfare and Treatment of Animals. Chronic colitis was achieved by transferring CD4+CD45RBhigh T cells into immunodeficient female SCID mice (6/8 weeks old). After confirming the development of pathology using endoscopy and MRI, animals were treated therapeutically with the RIP1 inhibitor GSK547A (50 mg/kg twice a day PO) or vehicle (0.5% hydroxypropyl methylcellulose in water). The severity of colitis was measured 5 weeks after cell transfer both macro- and microscopically. Mucosal damage by endoscopy was also assessed. RT-PCR and MSD analysis were performed to detect colon cytokine/chemokine and calprotectin levels. SAA in the plasma was measured using ELISA. Results We found that treatment with GSK547 significantly ameliorated experimental T-cell-dependent colitis in mice. GSK547A-treated mice displayed decreased weight loss, colon density (ratio weight/length), macroscopic disease activity index, colon thickness compared with vehicle-treated animals. Mucosal damage, assessed by endoscopy and histopathology, was also reduced following treatment with RIP1 kinase inhibitor. In addition, GSK547A reduced the expression of cytokines in the colon (TNF-α, IL-17A, INF-γ, IL-6 and MCP-1), both at a protein and RNA level. Relevant translational biomarkers such as SAA in plasma and RNA calprotectin in the colon were also decreased in the GSK547A treated group when compared with vehicle. Conclusion Our results suggest that RIP1K inhibition is a strong protective factor with anti-inflammatory potential in the progression of chronic colitis, when applied to the translationally relevant T-cell transfer model of colitis. These findings suggest the potential application in the management of inflammatory bowel disease and support the ongoing clinical program evaluating the effect of RIP1 kinase inhibition in UC patients.

Клиническое значение цитокиновых маркеров при различном течении внебольничной пневмонии у детей

Principles of the diagnosis and treatment of community-acquired pneumonia (CAP) in children were developed and clearly formulated long ago. Nevertheless, clinicians often encounter the problem of pulmonary and pleural complications of CAP, which is challenging in terms of the choice of initial therapy, since the first symptoms of uncomplicated and complicated pneumonia are often similar. Therefore, the search for early markers of complicated CAP in children is highly important. Objective. To assess prognostic values of spontaneous and mitogen-induced cytokine production in children with CAP. Patients and methods. We have performed comprehensive examination of 108 children with CAP. Eighty-four of them had uncomplicated CAP, whereas 24 children had CAP complicated by pleurisy. We measured spontaneous and induced production of the following cytokines upon patient admission to hospital: interleukin-1 (IL-1), interleukin-17 (IL-17), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), vascular endothelial growth factor (VEGF), and monocyte chemoattractant protein-1 (MCP-1). To measure induced cytokine production, we stimulated peripheral blood lymphocytes by S. рneumonае (serotype 7, 11; strains 7C and 11AD). The level of cytokines was evaluated using the enzyme-linked immunosorbent assay (Vektor-BEST, Novosibirsk, Russia). Results. We found that in children with uncomplicated CAP, induction of immunocompetent blood cells (IBCs) led to increased secretion of first-generation cytokines, including IL-1, TNF-α, and IFN-γ, whereas IBCs of patients with complicated CAP primarily produced second-generation cytokines, including VEGF, МРС-1, and IL-17. Conclusion. The observed differences in spontaneous and mitogen-induced cytokine production between children with and without CAP complications suggest that these parameters can be considered as promising prognostic markers for complicated CAP in children. The proposed method can be used in pediatric practice to predict the development of complications in children with CAP. Key words: children, community-acquired pneumonia, cytokines