Content Of Nitric Oxide Metabolites Research Articles

<i>NOS2</i>, <i>NOS3</i>, <i>SONE</i> gene transcripts levels in peripheral blood leukocytes and their relationship with markers of endothelial dysfunction in hypertension

Objective. The aim of the study was to evaluate the level of expression of the NOS2, NOS3, SONE genes in peripheral blood leukocytes (PBL) of patients with hypertension (HTN) and to study the relationship between the level of transcripts of these genes and the content of nitric oxide metabolites and markers of endothelial dysfunction.Design and methods. The study included healthy people (25 people) and patients with HTN (stages I–II) before prescribing antihypertensive drugs (15 people) and taking cardioselective β-adrenergic receptor blockers for more than a year (metoprolol (25 mg per day) or bisoprolol (5–10 mg per day)) (20 people). The level of gene transcripts was assessed by real-time polymerase chain reaction (PCR). The level of nitric oxide metabolites was determined by the colorimetric method using the Griess reagent. The content of asymmetric dimethylarginine (ADMA), soluble forms of vascular cell adhesion molecule (sVCAM), and intercellular adhesion molecule (sICAM) in blood plasma was determined by ELISA. The content of malondialdehyde (MDA) in blood plasma was determined spectrophotometrically by color reaction with thiobarbituric acid. Statistical processing of the results was carried out using the Statgraphics Centurion XVI software package (version 16.1.11).Results. The level of nitric oxide metabolites in the blood plasma of HTN patients without antihypertensive therapy was 2,1 times higher than in healthy individuals (p = 0,001) and 1,7 times higher than in patients with HTN taking metoprolol or bisoprolol (p = 0,002). The relative content of mRNA of the NOS3 gene in PBL of individuals included in the study did not differ (p > 0,05). The level of NOS2 gene transcripts in PBL of HTN patients before the prescription of antihypertensive drugs exceeded that in healthy individuals (p = 0,0009) and in HTN patients taking metoprolol or bisoprolol (p = 0,0002). The number of SONE transcripts in the PBL of HTN patients was higher than in people with normal blood pressure (p < 0,00001 when comparing patients before the prescription of antihypertensive therapy and individuals from the control group; p = 0,04 when comparing patients with HTN taking antihypertensive drugs and normotensive subjects). The content of MDA, ADMA, sVCAM was higher in the plasma of HTN patients without antihypertensive therapy compared with people from the control group (p = 0,005, 0,003, 0,039, respectively) and patients taking metoprolol or bisoprolol (p = 0,0006, 0,019, 0,016, respectively). The content of nitric oxide metabolites positively correlated with NOS2, SONE, VCAM1 mRNA level in PBL, the content of MDA and ADMA in blood plasma (p < 0,05). A positive correlation was found between the concentration of MDA and ADMA in plasma (p = 0,03).Conclusions. An increase in the level of nitric oxide metabolites in HTN is associated with an increase in the transcriptional activity of the NOS2 gene, a disturbance of the redox balance of the body, and the development of endothelial dysfunction. The SONE gene is probably involved in the modulation of nitric oxide levels in HTN not only as an antisense transcript that destabilizes the mRNA of the NOS3 gene in vascular endothelial cells, but also indirectly, namely, through the regulation of homeostasis of immune system cells through autophagy.

Electrochemical measurements of the levels of nitric oxide metabolites in the blood serum

Background: Sepsis is a serious clinical condition caused by a dysregulated immune response to infection resulting in multiple organ failure. In the pathogenesis of sepsis, especially that of septic shock, great importance is given to the endothelial marker of vascular regulation, nitric oxide (NO). In septic shock, dysregulation of the vascular tone plays a key role in the development of hypotension. Therefore, the control of the level of nitric oxide and its stable metabolites in critically ill patients is a very important task. Aim: the aim of this study was to evaluate the potential of the electrochemical nitrite detection in the patients blood serum. Methods: The levels of nitric oxide stable metabolites in the blood serum of healthy individuals (n=20) and septic patients (n=25) were studied by the electrochemical method using a composite electrode and by the spectrophotometric method using the Griess reagent. Results: The data in the groups of healthy people and patients with sepsis differ significantly (p 0.00001) both when measured using electrochemical and spectrophotometric methods. The median value of the current response in healthy people was 0.41 A (0.33; 0.55), and the total content of nitric oxide metabolites (NOx) was 26.8 mol/L (20.8; 31.0), while in patients with sepsis, these values were 0.79 A (0.61; 1.28) and 38.89 mol/L (29.64; 57.45), respectively. A negative correlation was found between the data obtained for practically healthy persons (r=-0.696, p=0.0007). Conclusion: The obtained results allow us to conclude that the nitrite measurement in the blood serum by amperometry using a composite electrode is promising as a diagnostic technique.

ВЫРАЖЕННОСТЬ ЭНДОТЕЛИАЛЬНОЙ ДИСФУНКЦИИ КАК ОБЪЕКТИВНЫЙ КРИТЕРИЙ СТЕПЕНИ ТЯЖЕСТИ ПРЕЭКЛАМПСИИ

Preeclampsia remains one of the most significant obstetric problems due to its high prevalence, high incidence of maternal and perinatal morbidity and mortality, as well as the need for significant economic costs for the management and delivery of pregnant women with this pathology. In recent years, a number of authors have proven the most important role of vascular endothelial dysfunction in initiating disorders of vascular tone and blood coagulation potential, which underlie the formation of multiple organ failure in pregnant women with preeclampsia. The aim of the study was to establish new objective criteria for assessing the severity of preeclampsia based on the study of the functional state of the vascular wall by various methods, both invasive and non-invasive. 118 women with 28-37 weeks of pregnancy were examined. The 1st observation group consisted of 58 pregnant women with moderate preeclampsia, the 2nd observation group included 20 pregnant women with severe preeclampsia. The control group consisted of 40 women with a physiological course of pregnancy. The functional activity of the vascular wall was assessed by the level of endothelin-1, thrombospondin, intercellular adhesion molecules and nitric oxide metabolites in the blood by enzyme immunoassay. In patients with moderate preeclampsia, the content of nitric oxide metabolites significantly decreased, and the content of endothelin-1, thrombospondin, and intercellular adhesion molecules increased relative to patients in the control group. These changes were significantly more pronounced in patients with severe preeclampsia. At the same time, to study the function of the endothelium in pregnant women with preeclampsia, we used a non-invasive method - an ultrasonic cuff test according to the Celermajer D.S. In patients with moderate preeclampsia, the degree of endothelium-dependent arterial expansion during the cuff test decreased by more than 2 times compared with the parameters of pregnant women in the control group. In pregnant women with severe preeclampsia of the group, an even more pronounced decrease in the degree of dilatation of the brachial artery was noted. Thus, the determination of markers of endothelial dysfunction in the blood, as well as the use of an ultrasonic cuff test, made it possible to identify violations of the vasomotor function in pregnant women with preeclampsia, which correlate with the severity of preeclampsia. The use of an ultrasonic cuff test and the determination of endothelial dysfunction markers can be recommended as additional objective diagnostic criteria for the severity of preeclampsia, the transition from moderate preeclampsia to severe preeclampsia.

Intensity of free-radical reactions in metastasizing cervical cancer.

e17508 Background: Cervical cancer (CC) is one of the main causes of death among patients with gynecological tumors. As a rule, patients die from relapses and metastases which rates reach 30-45%. According to research data, treatment of patients with distant metastases is ineffective, and half of them die within 9.7 months, while the average survival is 15.9±0.26 months. The development of many pathological processes is considered to be associated with an increase in free-radical reactions leading to oxidative damage to various biomolecules. The purpose of the study was to analyze some characteristics of the free-radical oxidation and antioxidant defence in metastasizing CC. Methods: The study included 56 patients aged 29-73 years with stage IIB-IV CC after antitumor treatment. The main group – 27 patients who developed metastases within 4 months to 8 years after the CC diagnosis; controls – 29 patients with non-metastatic CC; donors – 19 healthy women aged 27-61 years. The accumulation of carbonyl derivatives in blood plasma proteins was detected in the reaction with 2, 4-dinitrophenylhydrazine. The induced oxidative modification of proteins was stimulated with Fenton's reagent. Free-radical processes was evaluated by the intensity of peroxide-induced luminol-dependent plasma chemiluminescence and the content of nitric oxide metabolites; the intensity of lipid peroxidation - by the content of malondialdehyde (MDA); the activity of catalase and ceruloplasmin was also studied. Results: CC progression was accompanied by increasing lipid peroxidation and spontaneous oxidation of blood plasma proteins. MDA levels in patients with metastases increased by 57.8% compared to donors and by 34.3% compared to patients without metastases (p < 0.05). The concentration of 2, 4-dinitrophenylhydrozones increased on average by 4 times compared to donors and by almost 2 times compared to patients without metastases. Patients with metastasizing CC demonstrated elevated levels of products of the interaction of nitric oxide and its derivatives with proteins and peptides - 3-nitrotyrosine and nitrosoglutathione, compared to both donors (by 31.4% and 55.3%) and patients in remission (by 38.1% and 34.5%). Chemiluminescence activity increased by 54.5% compared to donors (p < 0.05) and by 93% (p < 0.01) compared to controls. Catalase activity in the blood plasma of patients with metastases increased by 54.1% compared to donors, but was lower than the values in the control group (by 22.1%). Ceruloplasmin activity was increased only in patients without metastases (by 33%). Conclusions: The process of CC metastasis is accompanied by a greater intensity of oxidative processes of both proteins and lipids, as well as depletion of the adaptive capabilities of the body's antioxidant system.

Формування адаптивних реакцій у Ceratodon purpureus (Hedw.) Brid. за участі хлорофілази та метаболітів оксиду азоту

Changes in chlorophylase activity and the content of nitric oxide metabolites in adaptive reactions of gametophyte of the cosmopolitan moss Ceratodon purpureus (Hedw.) Brid under different temperature regime, water deficit, high insolation on the dump of the mine “Nadiia” of Chervonohrad mining district of Lviv region have been studied. It was established that unfavorable conditions for the existence of plants due to lack of moisture, high surface temperature of the substrate and high insolation are created in the summer months on the dumps of coal mines. Due to the degradation of the photosynthetic apparatus in stressful conditions of technogenic-disturbed areas in chloroplasts of C. purpureus, the total content of chlorophylls was 294–413 μg/g of dry matter mass. In the gametophyte of moss in the localities on the terrace of the dump with a high insolation of 100 thousand lux., it was determined higher chlorophylase activity in 1.3 and 2.2 times, compared with the base localities (55 thousand lux.) and the top localities (70 thousand lux.), respectively. Under the influence of significant water stress, which was induced by a 10% solution of polyethylene glycol, a decrease in both the content of photosynthesis pigments and chlorophylase activity was determined compared to the control. The highest content of nitric oxide metabolites was determined on the terrace of dump due to their accumulation under the influence of significant insolation and temperature. The results of experimental studies of the effects of hypo-, hyperthermic and water stresses indicate the time-dependent changes in the content of nitric oxide metabolites in moss cells in the post-stress period. The increase in the NO2- content in the period of post-stress reaction indicates its important role in the adaptation of gametophyte of moss under the influence of abiotic stressors.

CONDITION OF ENDOTHELIAL FUNCTIONS OF VASCULAR WALL IN HYPERTENSIVE PATIENTS DURING DENTAL IMPLANTATION

Preoperative correction of periodontal vascular endothelial dysfunction in patients with essential hypertension (HD) can improve the efficiency of dental interventions and reduce the incidence of complications during the implantation process. The periodontal status and functional state of the vascular endothelium were examined in 60 patients with essential hypertension, aged 35 to 45, before and after implantation. The first, main group included 40 people with hypertension, who, in preparation for implantation, along with traditional dental treatment, were prescribed an antioxidant drug Mexidol. Patients of the main group were divided into two subgroups: in subgroup A, a solution of Mexidol was used in the form of applications on the mucous membrane of the gums; in subgroup B, the drug was administered both locally and per os. In the second group, in 20 patients with hypertension, surgical intervention was performed after the traditional complex of therapy. The control group consisted of 10 apparently healthy patients of the same age, with normal blood pressure. Endothelial dysfunction was diagnosed by the level of stable end metabolites of nitric oxide. During the initial examination of patients with hypertension, a significant decrease in the content of nitric oxide metabolites in the blood serum was revealed in comparison with the indicators of patients in the control group. After the course of the therapy, the indicators of nitric oxide in the serum of patients with hypertension increased, more significantly in those who received Mexidol, both locally and orally, which was accompanied by a clear improvement in periodontal status and the absence of complications in the postoperative period.

INFLUENCE OF CYTOKINES PRODUCTION, NITROGEN OXIDE METABOLITES AND LIPIDS EXCHANGE ON THE FORMATION OF EXTERNAL GENITAL ENDOMETRIOSIS STAGES IN PATIENTS OF REPRODUCTIVE AGE

The relevance of the study is due to the prevalence and increase in the incidence of endometriosis, especially in young patients, its influence on reproductive function, and the lack of reliable non-invasive diagnosis of the disease and its stages. There are no unified views on the etiology and multifactorial pathogenesis of this disease. This article is aimed at revealing the mechanisms of the formation of endometriosis stages basing on the study of the effect of cytokines production, nitric oxide metabolites and lipid metabolism. The main approach to study this problem is both to investigate systemic (blood serum) and local (peritoneal fluid) levels, their comparison. This allows comprehensive consideration of the pathogenesis of the disease and determination of the value of the mentioned biologically active factors depending on the stages of the disease. The article presents new data on the content of high- and low-density lipoproteins, cholesterol, transforming growth factor β1 and tumor necrosis factor α, the content of nitric oxide metabolites and their clinical interpretation is given according to the stages of the disease. The materials of the article are of practical value for researchers of endometriosis and practicing obstetrician-gynecologists.. Keywords: external genital endometriosis, lipoproteins, nitric oxide, transforming growth factor.

NITRIC OXIDE METABOLISM FEATURES UNDER CONDITIONS OF EXPERIMENTAL INFECTED RADIATION-INDUCED SKIN INJURIES DEVELOPMENT AND THEIR TREATMENT WITH PHOTODYNAMIC THERAPY

The aim: To follow-up nitric oxide content values in rat serum at the development of Staphylococcus aureus infected radiation skin injuries and their photodynamic therapy. Materials and methods: Eighty WAG male rats were studied in an experiment. Four groups were identified for evaluation. Group 1 included unaffected intact rats (n=20). Group 2 involved rats (n=20) with a modeled radiation-induced ulcer of the skin. The rats (n=20) with a modeled radiation-induced skin ulcer followed by infecting with Staphylococcus aureus were referred to group 3. Group 4 included rats (n=20) with Staphylococcus aureus infected radiation skin ulcer exposed to photodynamic therapy. Rats of groups 1-4 were sampled for biochemical blood examination on days 7, 14, 21, 30 and 45. Total nitric oxide metabolites (nitrites and nitrates) were measured according to V.A. Metelskaya et al. method. Results: Infectious agent (Staphylococcus aureus) present in skin ulcer impairs nitric oxide metabolism in rat blood serum that manifested in decreased total nitric oxide metabolites content on day 7, followed by its increase within days 14 to 45. While photodynamic therapy exposed on the Staphylococcus aureus infected radiation skin ulcer, total nitric oxide metabolites in blood serum had increased by day 7, but days 14 to 45 level was compliant with physiological norm. Conclusions: Infecting radiation skin ulcers with Staphylococcus aureus causes impaired nitric oxide metabolism, while photodynamic therapy helps to normalize the metabolism of the above-mentioned chemical compound that can improve healing of radiation skin ulcers.

Preventive Effects of Grape Extract on Ischemia/Reperfusion-Induced Acute Kidney Injury in Mice.

Since grape extract (GE) contains oligomeric proanthocyanidins and numerous polyphenols, dietary GE supplements may exert protective effects against various diseases. The present study investigated the pharmacological effects of GE derived from Chardonnay in vitro and in vivo. GE (100 µg/mL) completely inhibited tumor necrosis factor-α-induced endothelin-1, monocyte chemoattractant protein-1, interleukin-1β, and intercellular adhesion molecule-1 gene expression in cultured endothelial cells. GE also strongly stimulated the phosphatidylinositol 3-kinase (PI3K)/Akt/endothelial nitric oxide synthase (eNOS) pathway. In the in vivo study, the effects of GE on ischemic acute kidney injury (AKI) were examined using male C57bl/6J wild-type and eNOS-/- mice. Right nephrectomized mice were exposed to 45 min of ischemia in the left kidney and this was followed by reperfusion. Although renal functional parameters in AKI mice significantly increased 48 h after reperfusion, the administration of GE (0.1 and 1 mg/kg, intravenous (i.v.)) 5 min before ischemia dose-dependently improved post-ischemic renal dysfunction in wild-type mice. Renal histopathological studies on AKI mice revealed tubular necrosis, proteinaceous casts in tubuli, and medullary congestion. The administration of GE ameliorated this damage in wild-type mice, but not in eNOS-/- mice. Furthermore, GE significantly restored decreases in the renal nitric oxide metabolite content due to ischemia in wild-type mice, but not in eNOS-/- mice. Thus, eNOS is closely involved in the renoprotective effects of GE, strongly suggesting that GE supplements are useful as a prophylactic treatment for the development of ischemic AKI.

P2686Endothelial reaction mediates enhanced external counterpulsation effects in post-MI cardiac rehabilitation

Abstract Introduction The mechanisms mediating benefits of enhanced external counterpulsation (EECP) are disputable since the origin of the method. One of the possible explanations presumes increased diastolic flow leads to changes of endothelial function and plays a crucial role in alleviating the symptoms of angina and heart failure; the exact mechanisms of action are still under investigation, though the endothelial hypothesis possesses strong evidence. Purpose To reveal the impact of repeated EECP on endothelial function of patients undergoing cardiac rehabilitation program after myocardial infarction (MI) and to identify possible clinical implications of these changes. Methods The randomized sham-controlled study included 46 patients with coronary artery disease who suffered from ST-elevation MI (STEMI) 4–8 weeks prior to enrollment in 2017–2018. All patients had undergone primary PCI due to STEMI in 2–24 hours after the event. Echo parameters, exercise tolerance and the levels of ADMA and nitric oxide (NO) metabolites (total content, nitrite and nitrate) in plasma were determined at the enrollment stage and at three month follow-up. Patients were randomized and given 35 h of active counterpulsation (main group, n=24) or inactive counterpulsation (control group, n=22) over a six-week period after enrollment. Results The analysis showed no significant difference between groups at baseline: mean ejection fraction was 46% in the main group and 45% in the control group, no difference in exercise tolerance. The mean characteristics of endothelial function at baseline were similar in both main and control groups: total NO metabolites (17.3±2.14 μmol/L vs. 19.6±3.31, p=0. 42), nitrite level (0.76±0.17 μmol/L vs. 0.75±0.21, p=0. 36), ADMA level (0.78±0.15 μmol/L vs. 0.81±0.20, p=0. 22). At three month follow up 41,7% of patients of the main group showed improvement of one class of NYHA classification compared to 22,7% in the control group (U=122.3, p=0.012), agreed with improved exercise tolerance: average +2.8 MET in main group in opposite to only +0.8 MET in the controls (U=104, p=0.007). Control group demonstrated no significant dynamics of NO metabolites concentration comparing to baseline values, while in the main group there was a significant decrease of ADMA – 0.57±0.11 μmol/L (Z=r=−2.46, p=0.013) and increase of nitrite level – 1,12±0. 18 μmol/L (Z=r=−2.57, p=0.011), with no changes in total NO metabolites content. The further analysis demonstrated strong correlation between exercise capacity gain and ADMA decrease (R=r=−0.52, p=0.008) or nitrite elevation (R=0.48, p=0.012), with ADMA/nitrite ratio being the strongest predictor of clinical improvement (R=0.68, p=0.002). Conclusion Active EECP improves exercise capacity and heart failre symptoms in patients undergoing post-MI rehabilitation, with those effects being associated with endothelial function enhancement.

Effect of Transcutaneous Electrical Stimulation of Nerves on Blood Pressure and Blood Content of Neuropeptide CGRP and Nitric Oxide in Hypertensive Rats with Metabolic Disturbances.

The development of arterial hypertension in male Wistar rats with fructose-induced metabolic syndrome (12.5% of fructose solution as the only drinking source for 10 weeks) along with impaired glucose tolerance and increased serum concentration of triglycerides and LPO products caused a decrease in the content of serum blood calcitonin gene-related peptide (CGRP). Low-frequency transcutaneous electrical nerve stimulation (1 mA, 2 Hz, 10 min daily for 2 weeks) performed in 8 weeks after the beginning of fructose treatment reduced systolic BP and serum concentration of triglycerides and LPO produces and improved glucose tolerance. After stimulation, CGRP content in rats maintained on fructose diet returned to normal values and the content of nitric oxide metabolites increased. We hypothesize that CGRP and nitric oxide are involved in mechanisms mediating the therapeutic effect of low-frequency transcutaneous electrical nerve stimulation on arterial hypertension developing in metabolic syndrome.

The Indices of Nitric Oxide System in Rats with Carrageenan-Induced Enterocolitis Combined with Diabetes Mellitus

AbstractBackground and aims:Diabetes mellitus (DM) represents a considerable public health issue, being one of the major causes of morbidity and mortality in the modern societies. Chronic hyperglycemia is accompanied by significant physiological, biochemical, and histological changes, e.g. development of oxidative stress that affects the motor activity of the intestine. This study aimed to evaluate the indices of nitric oxide (NO) system in blood serum and colon tissue supernatant of rats with carrageenan-induced enterocolitis combined with streptozotocin-induced diabetes.Material and methods:The total NOS activity was determined by monitoring the rate of conversion of L-arginine into citrulline. The total quantity of NO metabolites was assessed by evaluating their amount, which included nitrite ions that were initially present in the sample (NO2−) and nitrate ions reducted to nitrites (NO3−).Results:We found a significant increase in total NOS activity in colon tissue of all experimental groups vs. control animals (54.8, 30.6 and 79.2 % respectively). The total content of NO metabolites in colon tissue of all experimental groups also significantly increased (2.8, 1.9 and 3.4 times respectively) compared to the control animals.Conclusions:We observed activation of nitroxydergic process in blood serum and colon tissue of rats with carrageenan-induced enterocolitis. Nitroxydergic processes markedly intensified in rats with carrageenan-induced enterocolitis combined with diabetes mellitus.

Assessment of endothelial cell function and physiological microcirculatory reserve by video microscopy using a topical acetylcholine and nitroglycerin challenge

BackgroundAssessment of the microcirculation is a promising target for the hemodynamic management of critically ill patients. However, just as the sole reliance on macrocirculatory parameters, single static parameters of the microcirculation may not represent a sufficient guide. Our hypothesis was that by serial topical application of acetylcholine (ACH) and nitroglycerin (NG), the sublingual microcirculation can be challenged to determine its endothelial cell-dependent and smooth muscle-dependent physiological reserve capacity.MethodsIn 41 healthy subjects, sublingual capillary microscopy was performed before and after topical application of ACH and NG. Total vessel density (TVD) was assessed in parallel using manual computer-assisted image analysis as well as a fully automated analysis pathway utilizing a newly developed computer algorithm. Flow velocity was assessed using space-time diagrams of the venules as well as the algorithm-based calculation of an average perfused speed indicator (APSI).ResultsNo change in all measured parameters was detected after sublingual topical application of ACH. Sublingual topical application of NG however led to an increase in TVD, space-time diagram-derived venular flow velocity and APSI. No difference was detected in heart rate, blood pressure, and cardiac output as measured by echocardiography, as well as in plasma nitric oxide metabolite content before and after the topical application of ACH and NG.ConclusionsIn healthy subjects, the sublingual microcirculatory physiological reserve can be assessed non-invasively by topical application of nitroglycerin without affecting systemic circulation.

Біохімічні зміни пародонта в щурів на тлі дії тютюнового диму

Introduction. In our days, smoking is one of the most important risk factors for inflammatory diseases. There is a lot of scientific data, devoted to research the effects of smoking on periodontal status. However, some aspects such as the status of free-radical oxidation, is left without proper attention.The aim of the study – to investigate experimentally the indices of free radical oxidation in periodontal tissues on the background of smoking.Methods of the research. Experimental studies were performed on 20 adult male albino rats weighing 160-180 g. Animals were divided into 2 groups: control (10 animals) and experimental (10 animals). Modeling of passive smoking was carried out by placing rats in a specially designed chamber in which tobacco smoke was distributed. The duration of the experiment was 21 days. The concentration of 8-isoprostane was determined by ELISA method using a set of "8-isoprostane ELISA» company «USBiological» (USA). Quantitative assessment of the content of metabolites of nitrogen (II) oxide was carried out by determining their amount. The content of reduced glutathione, superoxide dismutase and catalase activity in periodontal tissue homogenate were determined by conventional methods.Results and Discussion. Concentration of 8-isoprostane in supernatant of periodontal homogenate in rats of control group was (3.94±0.18) pg/ml. In experimental animals this index was significantly increased in 2.9 times. This indicates that despite the action of tobacco smoke in periodontal tissues oxidative stress develops, characterized by the increasing intensity of the production of reactive oxygen species. On 22 day of the experiment, we have observed a significant decrease in superoxide dismutase activity in periodontal tissue homogenate by 40.9 % and catalase activity in 2.4 times compared to the control group. It is known that SH-groups (such as cysteine and methionine fragments of protein molecules) are most easily oxidized by reactive oxygen species to form a reversible and irreversible modifications. On 22 day of the experiment, we have observed a significant decrease in glutathione content in periodontal tissue homogenate by 24.4 % compared to the control group. The content of NO metabolites in supernatant of periodontal homogenate in rats of experimental group was significantly decreased in 2.1 times. The powerful factors that inactivate NO, are free radicals, among them – superoxide radical. Its interaction with NO leads to formation of peroxynitrite that as a strong oxidant has high degree of cytotoxicity.Conclusions. In rats on the background of tobacco smoke in periodontal tissues significantly increases the generation of reactive oxygen species, reduces superoxide dismutase, and catalase activity and glutathione content, indicating a marked imbalance of free radical homeostasis. On the background of tobacco smoke in periodontal tissues occurs inhibition of nitroxydergic processes through significant reduction of nitric oxide metabolites content, which reduces the protective antibacterial potential.

Level of Nitrogen Oxide and Indicators of Oxidative-Antioxidative Status in the Gingival Tissues, Blood Plasma in Experimental Chronic Gastritis and Duodenitis and Drug Correction

model of chronic gastritis and duodenitis was conducted in 48 immature Wistar rats both gender by intragastric injection of medical bile. Control group consisted of 10 animals. After approximation model of the gastritis and duodenitis rats were divided into five subgroups: the 1st subgroup (before treatment, 9 rats), subgroup 2 (10 animals) received the antioxidant, subgroup 3 (11 animals) – received NO donator, subgroup 4 (10 animals) - received an antioxidant and NO donator, subgroup 5 (8 animals) – received the antioxidant, NO donator and calcium carbonate. In rats with gastritis and duodenitis was observed redistribution of stable metabolites of nitric oxide: decreasing level in the gingival homogenates and increasing in the blood plasma. Level of malondialdehyde and aldehyddehydrogenase in the gingival tissue increased, and in the blood plasma decreased, on a background of catalase activation; content of ketone phenilhydrazone remained in the level of control group. Decreasing markers of oxidation lipids and proteins in the blood plasma, on a background of increasing levels of catalase and nitric oxide metabolites (in 14 times), indicates about a key role of NO in the antioxidant protection of organism in a case of disease.Application of antioxidant could not impact on the indicators of oxidation lipids and proteins. It had been shown decreasing content of the nitric oxide metabolites in the gingival tissues and blood plasma. Drug correction with donator of nitric oxide have to increase markers of oxidation lipids and proteins in the gingival tissues, on a background of high catalase activity and low levels of nitric oxide metabolites. In the blood plasma content of nitric oxide metabolites was higher, which indicated about an active inflammatory process in the stomach and duodenum mucosa. Indicators of the protein molecules fragmentation and malondialdehyde were not differ from those before treatment. Level of catalase was in two times higher, than in the control group, but level of superoxide dismutase was decreased. Complex of antioxidant and donator of nitric oxide helped to stabilise the indicators of oxidation lipids and proteins, although level of nitric oxide metabolites in the gingiva was significantly decreased. Application complex from the antioxidant, donator of nitric oxide and calcium carbonate at the treatment of chronic gastritis and duodenitis restored an oxidative-antioxidative status in the blood plasma and gingival tissues, increasing the production of nitric oxide to a level in the control group.

LEVELS OF NITRIC OXIDE METABOLITES IN RATS WITH HEPATOPULMONARY SYNDROME

<p>Background. System of nitric oxide (NO), which consists of NO, and its metabolites, is very important for<br />various biological processes. NO is signalling molecules and mediators of intracellular and intercellular interaction<br />that causes relaxation of smooth muscles of blood vessel walls, inhibits platelet aggregation and their adherence,<br />is involved in the transmission of nerve impulses, cell proliferation.<br />Objective. The aim of our research was to study the content of nitric oxide metabolites in blood serum and<br />bronchoalveolar lavage, to substantiate their role in pathogenesis of hepatopulmonary syndrome in<br />experiment.<br />Methods. The experiments were performed on 56 outbread male rats, 180-220 g in weight. The first experimental<br />model of hepatopulmonary syndrome (HPS) was made by imposition of double ligature on common<br />bile duct and its further dissection with a scalpel. The second experimental HPS model was made by 8-week intragastric<br />administration of oil solution CCl4 (400 g per 1 L), 0.5 ml per 100 g of body weight on the first day of<br />the experiment, 0.3 ml per 100 g on the third day of the experiment and then every third day until the end of the<br />experiment 0.3 ml per 100 g. A mixture of corn flour, lard and cholesterol and alcohol solution was added to the<br />standard diet of the rats.<br />Results. The total content of nitric oxide metabolites in blood serum of the rats of the experimental group<br />No.1 (on the 31st day after the common bile duct ligation) was significantly increased in 3.9 times (p1<0,001) if<br />compared with the control group №1. In the rats of the 2nd experimental group (with carbon tetrachloride induced<br />cirrhosis) the total content of nitric oxide metabolites in blood serum also significantly increased in 3.1 times<br />(p1<0,001). Comparison of nitric oxide metabolites content in blood serum and bronchoalveolar lavage, which<br />directly indicated about the processes in lung tissue, was great importance.<br />Conclusions. So, in rats with experimental hepatopulmonary syndrome activation of nitroxydergic process<br />by significant increase in nitric oxide metabolites in blood serum and bronchoalveolar lavage took place.<br />KEYWORDS: hepatopulmonary syndrome, nitric oxide metabolites.</p>

The stress effects of a single injection of isotonic saline solution: systemic (blood) and central (frontal cortex and dorsal and ventral hippocampus)

In experiments with animals, a group that is injected with the vehicle in which a drug of interest is dissolved is often used as a control. However, even a single injection of a vehicle is a stressor, i.e., “treatment stress,” which may significantly affect some stress-sensitive indices. In the present study, we report some data on the effects of a single intraperitoneal injection of isotonic saline solution on the contents of corticosterone, nitric oxide metabolites, and oxidative capacity, as well as on the expression of proteins and mRNAs of proinflammatory cytokines in the blood and brain regions of rats within one day after the injection as compared to intact animals. At the early time points after the injection, corticosterone contents were substantially elevated in the blood and ventral hippocampus. The content of nitric oxide metabolites decreased in the blood and remained stably low within 2–24 h after the injection. The injection did not affect the contents of proinflammatory cytokines in the blood; however, early after the injection the expression of IL-1s mRNA decreased in the ventral hippocampus and frontal cortex, whereas 24 h after this treatment, the expression of TNF-a mRNA increased by a factor of 4 in the frontal cortex. Thus, a single injection of isotonic saline solution had a clear stress-producing effect, which was observed at the systemic level and in stress-sensitive brain regions. The strength of this stressful event was sufficient to activate the hypothalamus–pituitary–adrenal axis but not sufficient to induce a significant inflammatory response. The frontal cortex was most sensitive to this treatment; the alterations in the ventral hippocampus were less expressed, whereas the dorsal hippocampus was most stress resistant. Our data show that it is important to consider and thoroughly analyze the effects of “treatment stress” in experiments using injections of biologically active substances.

HOE-140, an antagonist of B2 receptor, protects against memory deficits and brain damage induced by moderate lateral fluid percussion injury in mice

There are evidences indicating the role of kinins in pathophysiology of traumatic brain injury, but little is known about their action on memory deficits. Our aim was to establish the role of bradykinin receptors B₁ (B₁R) and B₂ (B₂R) on the behavioral, biochemical, and histologic features elicited by moderate lateral fluid percussion injury (mLFPI) in mice. The role of kinin B₁ and B₂ receptors in brain damage, neuromotor, and cognitive deficits induced by mLFPI, was evaluated by means of subcutaneous injection of B₂R antagonist (HOE-140; 1 or 10 nmol/kg) or B₁R antagonist (des-Arg9-[Leu8]-bradykinin (DAL-Bk; 1 or 10 nmol/kg) 30 min and 24 h after brain injury. Brain damage was evaluated in the cortex, being considered as lesion volume, inflammatory, and oxidative damage. The open field and elevated plus maze tests were performed to exclude the nonspecific effects on object recognition memory test. Our data revealed that HOE-140 (10 nmol/kg) protected against memory impairment. This treatment attenuated the brain edema, interleukin-1β, tumor necrosis factor-α, and nitric oxide metabolites content elicited by mLFPI. Accordingly, HOE-140 administration protected against the increase of nicotinamide adenine dinucleotide phosphate oxidase activity, thiobarbituric-acid-reactive species, protein carbonylation generation, and Na⁺ K⁺ ATPase inhibition induced by trauma. Histologic analysis showed that HOE-140 reduced lesion volume when analyzed 7 days after brain injury. This study suggests the involvement of the B₂ receptor in memory deficits and brain damage caused by mLFPI in mice.

The effect of NADPH-oxidase inhibitor apocynin on cognitive impairment induced by moderate lateral fluid percussion injury: Role of inflammatory and oxidative brain damage

Traumatic brain injury (TBI) is a devastating disease that commonly causes persistent mental disturbances and cognitive deficits. Although studies have indicated that overproduction of free radicals, especially superoxide (O2-) derived from nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is a common underlying mechanism of pathophysiology of TBI, little information is available regarding the role of apocynin, an NADPH oxidase inhibitor, in neurological consequences of TBI. Therefore, the present study evaluated the therapeutic potential of apocynin for treatment of inflammatory and oxidative damage, in addition to determining its action on neuromotor and memory impairments caused by moderate fluid percussion injury in mice (mLFPI). Statistical analysis revealed that apocynin (5mg/kg), when injected subcutaneously (s.c.) 30min and 24h after injury, had no effect on neuromotor deficit and brain edema, however it provided protection against mLFPI-induced object recognition memory impairment 7days after neuronal injury. The same treatment protected against mLFPI-induced IL-1β, TNF-α, nitric oxide metabolite content (NOx) 3 and 24h after neuronal injury. Moreover, apocynin treatment reduced oxidative damage (protein carbonyl, lipoperoxidation) and was effective against mLFPI-induced Na+, K+-ATPase activity inhibition. The present results were accompanied by effective reduction in lesion volume when analyzed 7days after neuronal injury. These data suggest that superoxide (O2-) derived from NADPH oxidase can contribute significantly to cognitive impairment, and that the post injury treatment with specific NADPH oxidase inhibitors, such as apocynin, may provide a new therapeutic approach to the control of neurological disabilities induced by TBI.

Purple grape juices prevent pentylenetetrazol-induced oxidative damage in the liver and serum of Wistar rats

Oxidative damages in hepatocytes may be caused by epilepsy and/or anticonvulsant drugs. Epilepsy is one of the most common neurological disorders, characterized by recurrent seizures, which may increase the content of reactive oxygen species. Organic and conventional grape juices are rich in polyphenols, compounds with important antioxidant activity. It is hypothesized that organic and conventional purple grape juices may have protective effect against oxidative damage induced by pentylenetetrazole (PTZ) (a standard convulsant drug) in the liver and serum of Wistar rats. Animals (n = 16 in each group) received, by gavage, saline, organic grape juice or conventional grape juice (10 μL/g of body weight) for 17 days. Subsequently, half of the rats in each group received PTZ (60 mg/kg). After 30 minutes, the animals were euthanized by decapitation. Liver and blood samples were isolated to evaluate oxidative parameters (lipid and protein oxidation, nitric oxide metabolite content, antioxidant defenses, and protein sulfhydryl content). The results of this study showed that although organic juice contains higher polyphenol content than conventional juice, both juices conferred protection against lipid and protein oxidative damage and limited the increase in PTZ-induced nitric oxide metabolite content in the liver and serum. In addition, both juices inhibited the PTZ-induced reduction in enzymatic antioxidant defenses (superoxide dismutase and catalase activities) and sulfhydryl protein content in the liver and serum. In summary, both organic and conventional grape juices were able to reduce oxidative damage induced by PTZ in the liver and serum of Wistar rats.