[Progesterone - regulator of T-lymphocytes cytotoxicity].

Progesterone plays important the role in the regulation of the immune system during pregnancy. We examined the effect of progesterone on the cytotoxic activity of T-lymphocytes in a model system Jurkat cells. Jurkat cells were stimulated with 50 µg/ml of phytohemaglutinin A (PHA) at 370C for 5 minutes. Then, PHA was removed by centrifugation, the cells were washed and cultured for 24 hours alone or with progesterone (added to the incubation medium of Jurkat cells at a concentration of 0.07 and 0.7 µl. Determined value of The parameters of mitochondrial membrane potential (by flow cytometry) and parameters of mitochondrial oxidative metabolism (the rate of generation of superoxide and peroxide radicals and activity of mitochondrial superoxide dismutase, catalase, glutatinperoxidase, the degree of nitrosylation of mitochondrial electron-transport proteins (by method of electron paramagnetic resonance and spectrophotometry) were determined. It was identified the lowering effect of high dose of progesterone (0.7µl) on the value of mitochondrial membrane potential (balancing percentage of cells with high and low values of mitochondrial potential and decreased intensity of oxidative stress in mitogen - activated Jurkat cells, which supports inhibition of their proliferation and differentiation activity.

ß2-adrenergic regulation of T lymphocites function (in vitro study).

The aim of our study was to establish the influence of ß2AR agonists and antagonists on Th1/Th2 subpopulation balance in intact and activated CD4+ T lymphocyte. Jurkat leukemic T cell line was used as a model for studying T cell activation conditions under the influence of ß2AR ligands. As follows from the results of our studies, after the influence of ß2AR agonist isoproterenol on intact Jurkat cells expression of IL-2 was not changed in comparison to control level. Under the PHA-stimulation level of IL-2 production in Jurkat cells increased significantly; isoproterenol caused decrease level of IL-2 expression in the PHA-stimulated Jurkat cells. Adding of ß2AR antagonist propranolol to the Jurkat cells pre-incubated with isoproterenol didn't change expression of IL-2. ß2AR antagonist propranolol induced slight increase of IL-2 expression in PHA-stimulated Jurkat cells pre-incubated with isoproterenol. Neither isoproterenol nor propranolol didn't change intensity of IL-10 expression in intact Jurkat cells. In the PHA-stimulated Jurkat cells level of IL-10 production decreased in comparison to control level. Isoproterenol induced sharp intensification of IL-10 expression in these cells. Propranolol prevented increase of IL-10 expression in the PHA-stimulated Jurkat cells pre-incubated with ß2AR agonist. It was concluded that ß2ARs in dose-dependent manner regulate cytokine profile in intact and mitogen activated CD4+ T lymphocyte and by this way induce dose-dependent alterations of lymphocyte proliferation and immune response. This indicated existence of a link among immune response and sympathetic nervous system activity.

[The role of neuroendocrine mediators in regulatory activity of T-cells].

The aim of the study was to establish the role of some neuroendocrine mediators (agonists and antagonists of ß-adrenergic receptors, progesterone) in regulating T-cells activity. Studies conducted on the culture of leukemiatransformed T-cells (Jurkat cells) (DSMZ-Deutshe Sammulung von Mikroorganismen und Zellkulturen (Germany)). Jurkat Cells (4 x 105 cells/ml) stimulated with 50 µg/ml fitogemaglutinin A (PHA) at 370С for 5 minutes and then incubated for 24 hours alone, or together with ß- adrenergic receptors agonist izopretonolom (at dose of 10-5 M, 10-6 M), an antagonist, propranolol (dose of 10-5 M, 10-6 M) and progesterone (dose 0,07 µl, 0,7 µl) added to the incubation medium. The viability of Jurkat cells was determined by MTT test. It was shown that ß-adlenoretseptors agonist, izoprotenol didn't affect the activity of mitochondrial dehydrogenases in intact and contributed to their low activation (12%) in the mitogen-activated Jurkat cells. ß-adrenergic receptors antagonist, propranolol, promotes a significant reduction in activity of mitochondrial dehydrogenases, and hence the viability of both intact (40-60%) and mitogen stimulated Jurkat cells (20-40%). Viability of intact Jurkat cells didn't change, and dose-dependently increased in PHA-stimulated Jurkat cells during progesterone exposure. It was concluded that viability of the T-cells and hence their functional activity is largely sensitive to the influence of the ß-adrenoceptor antagonists and progesterone. These data should be considered in the clinical application of appropriate drugs.

[Disorders of neurogenesis of cortical and subcortical structures in rat brain limbic system during fetal alcohol syndrome formation].

Disorders of neurogenesis of cortical and subcortical structures in rat brain limbic system were studied in the offspring of rats that received ethanol during pregnancy. The methods used included the staining of histological sections with cresyl violet, in vitro culture, and electron paramagnetic resonance. Prenatal alcohol intoxication was shown to induce the disturbances in proliferative activity of granular layer cells in the hippocampal dentate gyrus, neuron- and glioblast migration, enhancement of free NO and lipoperoxide production and cell death. This resulted in the changes in the number of neurons in cortical and subcortical structures of rat brain limbic system and in fetal alcohol syndrome formation.

[Investigation of metabolic paramagnetic centers in the blood of patients with prostate adenocarcinoma prior to and after the plastic orchectomy].

Paramagnetic centers in the blood of patients with prostate adenocarcinoma have been studied prior to and after plastic orchectomy by EPR. It has been shown that significant alterations in blood paramagnetic centers occur after the surgery. These alterations point to an improvement of antioxidant properties of the blood. Specifically, the content of the Fe(3+)-transferrin complexes increased, and the number of inactivated adrenoreceptors, MetHb, and Mn(2+)-containing complexes decreased. It was found that, along with improvement of the antioxidant properties of the blood, a significant decrease in the intensity of lipid peroxidation in the blood of the patients occurs.

[Study of toxicity of denture prosthetic appliance prothyl hot on the Jurkat cell model system].

The aim of our study was the investigation of toxicity of denture prosthetic appliance Prothyl Hot on the model system of Jurkat cell culture. As a result of our study it was revealed that denture prosthetic appliance Prothyl Hot: - hadn't manifested toxic effects the viability Jurkat cells (that reviled by stability of activity of mitochondrial dehydrogenises and mean of mitochondrial membrane potential; - didn't influence on the balance of pro-and antyinflammatory cytokines, expressed by Jurkat cells; - induces intensification of oxidative metabolism in Jurkat cells, which may be considered as compensatory reaction developing in the cells.

[Some considerations of immune metabolism at preeclampsia].

The goal of the work is to determine several aspects of abnormalities and a part of immune metabolism in preeclampsia. In peripheral blood serum of all pregnant women at 20th week of gestation relative content of T-lymphocyte subpopulations, active T-lymphocytes and natural killers (NK) were detected by the method of indirect immunofluorescence and in cyto-toxic test using monoclonal antibodies to CD4 and CD84, CD 16 (T-cell markers of HLA-DR antigen of second class of MHC (marker of active T-lymphocytes, "ICN Pharmaceutical", USA). According study results it can be concluded that at preeclampsia a content of NK cells in peripheral blood of pregnant women increases in the end of the second and early in the third trimesters on the background of significant decrease of general CD4 lymphocyte subpopulation. Received data allow to suppose that activation of natural killer cells (NK-cells) may be caused by the activation of cytotoxic Th-1 cells and their cytokines.

[Correction of oxidative stress in the rat brain cortical cellular culture with vitamines E and C].

The oxidative stress-induced alteration in concentration of oxygen, lipid and nitrogen free radicals in cultured brain cortex of the newborn albino rats and its correction with vitamins E and C was investigated. Modeling the oxidative stress was achieved with addition of H(2)O(2) into the nutrient medium. In order to prevent an oxidative stress-induced cytotoxic effect, concomitantly with H(2)O(2), vitamins E and C were added into the nutrient medium. Oxygen, lipid and nitrogen free radicals were registered by method Electron Paramagnetic Resonance (EPR) and spin-traps. The data, obtained in our study revealed decreasing of intensity of NO content and increasing of spin trapped superoxid- (O(2-)) and lipoperoxid-radicals (LOO.) in explants of brain cortex of the newborn albino rats cultured in oxidative stress conditions. These changes were attenuated following action of vitamins E and C (decreasing of intensity of EPR signals of O2- and LOO, increasing of free NO content).

[Pro- and antioxidant activity of bilirubin, EPR study].

The aim of the study was establishment of mechanisms of bilirubin oxidation and their involvement in the physiological and pathological processes in the living body (EPR study of photoradiated bilirubin). The photosensitized formation of free radical of bilirubin with g=2.003 and DeltaH=1.0 mTl, under action of the blue light with lambda(max)=450 nm by means of electronic spin resonance (ESR) was shown. Irradiation of sample in vacuum by visible light does not cause formation of free radicals. Irradiation of powder of bilirubin and also of its solution in chloroform leads to formation of the radical of bilirubin. The analysis of a spectrum (ESR) as powder also its solution in chloroform, that induced free radical belongs to bilirubin but not of solution was shown. Irradiation of a solution of bilirubin in chloroform causes absorption spectrum with lambda(max)=650 nm, characterized for absorption of solutions biliverdin in chloroform.

[The condition of oxidation-antioxidation balance of kidney and suprarenal tissue in experimental traumatic shock with white rats and the influence of Plaferon LB].

The aim of the work is the study of the oxidation-ant oxidation balance of kidney and suprarenal tissue and the influence on them of Plapherone LB preparation. The objects for experiment were 30 white rats which weigh was about 200gr. The traumatic shock was reproduced according to Kennon. White rats were divided into two groups. In 15 minutes after shock intraperitonealy were made: first group - 0,3 ml of physiologic solution, second group - Plapherone LB 0,06 mg on 0,3 mg of physiologic solution. Separately was studied control group of animals. The tissues were studied by the method of electronic paramagnet resonance on RE-1304 radiometer (Russia). The rough upset of oxidation-antioxidation balance was revealed. The use of Plapherone LB in this case promotes the optimization of these changes.

[Influence of antioxidants under polytrauma during traumatic sickness].

To study protective effect of Plapheron LB during grave traumatic sickness. Studied 94 patients with grave polytrauma, who were divided in to two groups of 47 patients. To I-st group was prescribed ordinary program of intensive therapy and to the II-nd group of patients additionally to the ordinary program of intensive therapy was made Plapheron LB in dose of 0.028 mg/kg two times a day sublingually during ten days. Grave condition of the patient was fixed by RTS scale and grave of shock by Skinner scale. Stated that each group was different in grave traumatic and shock conditions. Separately was studied group of health patients. The study was held by the method of electro-paramagnetic resonance of venous blood to admitted to hospital patients and on fifth day of intensive therapy. Noted: NO, FeS-NO, Fe(3+)-transpherin, Fe(2+) and Met-Hb. From immunology cytokin TNF-alpha was studied by the method of immunopherment. Results of study and discussion: Plapheron LB promotes decrease of the process of peroxide oxidation of lipids, increases activation of antioxidant ferments, stimulates production of nitric oxide and decreases the quantity of pyoinflammatory complication from those organs and systems which has no frequent effect of medical manipulation.

[Cell death in Jurkat cells induced by oxygen/nitrogen stress].

The maintenance of balance between lymphocyte proliferation and lymphocyte death is extremely important for normal functioning of the immune system. Considering essential role of nitrogen and oxygen radicals in functioning of immune competent cells, we studied the mechanisms of cell death induced by nitrogen/oxygen stress on the model of Jurkat cell line. We have observed that hyperproduction of reactive oxygen in Jurkat cells incubated with hydrogen peroxide contributes to the activation of membrane peroxidation, to a decrease in the intensity of apoptosis and its replacement by more severe mechanism of cell death - necrosis, which is obviously conditioned by a dramatic decrease in the intensity of energogenesis in mitochondria. In cells incubated with sodium nitroprusside moderate NO-induced inhibition of electron transport in oxidative chain and mitochondrial energogenesis and intensification of oxidative stress in Jurkat cells is accompanied with the activation of the cell death mechanisms- both apoptosis and necrosis.

[Investigation of oxidative stress-induced alterations in the rat brain cortical cellular culture and their correction with vitamines E and C].

The oxidative stress-induced changes and feasibility of prevention of these changes with an aid of vitamines E and C were investigated in cultured brain cortex of the newborn albino rats. Modeling the oxidative stress was achieved with addition of H(2)O(2) into the nutrient medium. In order to prevent an oxidative stress-induced cytotoxic effect, concomittantly with H(2)O(2), vitamines E and C were added into the nutrient medium. The data, obtained in our study revealed inhibition of the glial cells'- and axonal migration into the growth zone resulting from the oxidative stress. These changes were attenuated following action of vitamines E and C, which points at feasibility of their use against oxidative impact from the free radicals emerging during neurodegenerative disorders.

[Mechanisms of gamma-inducible death of Jurkat cells line].

Mechanisms of radio-inducible death of Jurkat cells were investigated. Human lymphoblastoid T-cell line Jurkat is widely established model for studying apoptosis mechanisms. The cell was radiated by "Teragam" (Czech Republic) by dose 2 g during 1 minute. After radiation cells were incubated at standard conditions during 24 hours. After gamma radiation in cell population amount of cells in gaplois (apoptotic G 0) stage was increased 8,2 folds, in diplois (G 0/G1) stage - by 17%, in synthetic (S) stage decreased by 35% and tetraploid (G2/M) stage by 73% in comparison to control group. It was revealed intensive production of free radicals of oxygen and nitric oxide and decreasing activity of antioxidant enzymes (superoxidismutasa, catalasa and glutathione peroxidase). Revealed dependence between intensification of apoptosis and radiation-induced arrest of cell cycle G2/M phase may be determined by excess amount of free oxygen and nitrogen radicals generated in Jurkat cells as a result of nondirect effects of low doses of gamma radiation.

[The role of endothelial dysfunction and Chlamydia pneumoniae infection in patients with ischemic stroke].

The research purposed to investigate the Chlamydia Pneumoniae infection in patients with ischemic stroke and to establish the correlation with elevated C. pneumonia antibody titers and endothelial dysfunction. 72 patients with acute ischemic stroke, aged 45 to 75 (45 male and 32 female) have been researched. Patients were grouped according to etiology of stroke (based on International classification TOAST). The blood was taken in 48 hours from stroke onset. ELISA method was applied to detect the antibodies against C. Pneumoniae. The intima-media thickness of the common carotid artery were assessed by B-mode ultrasonography (Acuson128XP/10) with a 7.5-MHz linear-array transducer. Flow-mediated dilatation (FMD) of the brachial artery were measured from high-resolution, 2-dimensional ultrasound images obtained by an ultrasound machine with a 7.5-MHz linear-array transducer. Free NO were examined by Electron Paramagnetic Resonance (EPR) method in blood. Control consisted with 15 healthy volunteers. Multivariate logistic regression showed a significant positive correlation between blood elevated titers of IgA and IgG of stroke patients and the intima-media thickness of the carotid artery (correlation coefficient by Pearson r = 0,31; p<0,05, r = 0,27; p<0,05). On multiple regression analysis, percent FMD showed a significant negative correlation with the intima-media thickness of the common carotid artery (r= -0,57, p<0,05), positive correlation between NO and the intima-media thickness of the carotid artery (r= 0,72, p<0,05). Association between antibody seropositivity for C. pneumoniae and increase of the IMT in the common carotid artery indicates persistent chronic infection in patients with ischemic stroke. Correlation between NO, an increase of the IMT in the common carotid and artery flow-mediated dilation of brachial artery indicates on the participation of NO in development of endothelial dysfunction and its significant role in pathogenesis of ischemic stroke. These findings support the concept that NO-inducible endothelial dysfunction is related to atherogenesis.

[The status of redox system in patients with Graves' disease].

The aim of our investigation was studying the redox processes, the establishment of state of blood oxidative and antioxidant systems in patients with Graves' Disease (GD). 19 with diffuse-toxic goiter have been investigated. Control group included 10 practically healthy subjects. The parameters of blood redox-system were investigated by electron-paramagnetic resonance method (EPR). In GD group hypothyroidism was diagnosed in 7 patients (36,8%), euthyroidism - in 7 patients (36,8%) and hypothyroidism - in 5 (26,3%). The value of ceruloplasmin in main group was significantly higher than in the control (18,6+/-1,3 mm/mg vs. 16,0+/-1,1 mm/mg, p<0,001); EPR-signal of Fe(3+)-transferrin in main group was also significantly higher (19,2+/-1,2 mm/mg vs. 22,0+/-0,9 mm/mg, p<0,001); values of nitric oxide in both groups were almost identical. EPR-signals of Mn(2+) ions significantly correlated with thyroid volume (r=0,5461, p=0,016), NO - with antibodies to thyroglobulin (r=0,5750, p=0,010). These changes demonstrate possible relationship between autoimmune diseases and redox-system.

[The role of oxidative stress in pathogenesis of GBS].

Axon degeneration accompanying its demielinization is a main course of neurological insufficiency typical for GBS. The mechanisms of axon degeneration, considered as the secondary result of serve inflammation are not established. We aimed to determine the role of oxidative metabolism in viral polyneuropathy pathogenesis. The activity of pro- and antioxidant systems of the body was studied by electron paramagnetic resonance (EPR) method. In blood and cerebrospinal fluid the intensive EPR signals of nitric oxide (NO), complexes of NO with nonhemic iron (HbNO), lypo- and superoxide radicals content noticeably increases, the signals of free Mn2+ and Fe2+ revealed, the activity of blood antioxidant enzymes, ceruloplasmin and katalasa increases (by 60%), superoxidedismitase's and glutation reductases activity decreases (by 20% and 70% correspondingly). It was considered, that inflammatory damage of nervous system induced by different infectious stimulus is initiated by activated immune cell proinflamatory agents (reactive oxygen and nitrogen species). Subsequently the oxidative stress, as result of accumulation of generators of reactive oxygen species, disordered intracellular metabolism products, contributes to axon demielinization and degeneration.

[Oxidative metabolism parameters in newborns with sepsis during the cerebral ischemia of different degrees].

Within the last decade neonatal sepsis remains one of the actual problems of medicine due to its high morbidity and lethality rates. Reference data clarify that in septicaemia pathogenesis a great importance is attributed to the cellular oxidation-reduction imbalance and oxide stress development. A redox imbalance developed during sepsis indicates to the cell's dysfunction. Also central nervous system is one of the first organs, which is damaged during the sepsis. The purpose of our research was the investigation of activity of pro- and antioxidant system in newborn babies with sepsis during the cerebral ischemia of different degrees. It was shown that during the newborns' sepsis the intensity of free-radical oxidation-reduction process was increased remarkably, antioxidant system activity decreased and intensive ESR-signal of HbNO MetHb was fixed in the blood. These changes caused to development of hypoxemia and deep cerebral ischemia.

Role of free nitrogen and oxygen radicals in the pathogenesis of lipopolysaccharide-induced endotoxemia.

We studied molecular mechanisms of changes in oxidative metabolism under conditions of experimental lipopolysaccharide-induced endotoxemia. Generation of reactive nitrogen and oxygen species in mice increased 18 h after treatment with lipopolysaccharide. These changes contributed to inactivation of enzymes and enzyme complexes (ribonucleotide reductase, NADH-ubiquinone oxidoreductase, and cytochrome c oxidase), dysfunction of the mitochondrial electron transport chain, and development of oxidative stress. Plaferon LB protected mice from the toxic effect of lipopolysaccharide.

[The role of oxidative stress in pathogenesis of obesity].

Recent findings on fat metabolism open new prospective for developing effective drugs that would focus on white adipose tissue as therapeutic target. The aim of our study was establishment of features of oxidative metabolism in adipose tissue in diet-inducible obesity (DIO) model in rats and under influence of citrus. By the result of our investigations it may be concluded, that adipose tissue plays important role in development of oxidative stress during obesity. In DIO model in adipose tissue it was revealed disorders of mitochondria electron transport chain, excess generation of reactive oxygen species and lipoperoxides, inactivation of superoqsiddismutasa, intensification of lipid peroxidation and development of oxidative stress. Citruses by the normalization of body lipid metabolism promotes normalization of mitochondrial electron transport, decrease of oxidative stress, restoration of adipocytes' metabolism.