Role of glutathione metabolism and glutathione-related antioxidant defense systems in hypertension.

The risk of developing chronic hypertension increases with age. Among others factors, increased oxidative stress is a well-recognized etiological factor for the development of hypertension. The co-occurrence of oxidative stress and hypertension may occur as a consequence of a decrease in antioxidant defense system activity or elevated reactive oxygen species generation. Glutathione is a major intracellular thiol-disulfide redox buffer that serves as a cofactor for many antioxidant enzymes. Glutathione-related parameters are altered in hypertension, suggesting that there is an association between the glutathione-related redox system and hypertension. In this review, we provide mechanistic explanations for how glutathione maintains blood pressure. More specifically, we discuss glutathione's role in combating oxidative stress and maintaining nitric oxide bioavailability via the formation of nitrosothiols and nitrosohemoglobin. Although impaired vasodilator responses are observed in S-nitrosothiol-deficient red blood cells, this potential hypertensive mechanism is currently overlooked in the literature. Here we fill in this gap by discussing the role of glutathione in nitric oxide metabolism and controlling blood pressure. We conclude that disturbances in glutathione metabolism might explain age-dependent increases in blood pressure.

Glutathione-related antioxidant defense system in elderly patients treated for hypertension.

The purpose of this study was to analyze glutathione antioxidant defense system in elderly patients treated for hypertension. Studies were carried out in the blood collected from 18 hypertensive and 15 age- and sex-matched controls, all subjects age over 60. Hypertensives were on their usual antihypertensive treatment at the time of blood collection. The concentration of glutathione (GSH) in whole blood and activities of glutathione peroxidase (GPx-1), glutathione transferase (GST), and glutathione reductase (GR) in erythrocytes were measured. The data from patients and controls were compared using independent-samples t test. P value of 0.05 and less was considered statistically significant. We observed increased glutathione-related antioxidant defense in treated hypertensive elderly patients (HT) when compared with healthy controls (C). Mean GSH concentration was significantly higher in HT when compared with C: 3.1 ± 0.29 and 2.6 ± 0.25 mmol/L, respectively, P < 0.001. Mean activity of GR was significantly higher in HT group if compared with C: 83.4 ± 15.25 U/g Hb versus 64.2 ± 8.26 U/g Hb, respectively, P < 0.001. Mean activity of GST was significantly higher in HT group compared with C: 3.0 ± 0.60 mmol CDNB-GSH/mgHb/min and 2.6 ± 0.36 mmol CDNB-GSH/mgHb/min, respectively, P < 0.05. No difference in GPx activity was observed between two groups. These results show that glutathione-related antioxidant defense system was enhanced in elderly hypertensive patients treated for their conditions. This suggests important role of glutathione system in blood pressure regulation. Alterations in concentration and activity of antioxidants observed during antihypertensive medication are likely to be related to the effect of the treatment on NO bioavailability.

Attitudes to brain death and organ procurement among university students and critical care physicians in poland.

PURPOSE: The practice of retrieving vital organs from brain-dead heart-beating donors is legally and medically accepted in Poland, but public beliefs and opinions regarding these matters have not been sufficiently explored. The purpose of this study was to evaluate the attitude of university students to the concepts of brain death and organ retrieval, compared with the attitude of critical care physicians. METHODS: The cohorts of 989 students and 139 physicians completed a questionnaire based on a survey instrument developed in an earlier reported study on Ohio residents. Participants assessed 3 scenarios: (1) brain death, (2) coma, and (3) vegetative state. More than 48% of students classified the patient from the brain death scenario as alive, and 51% of them were willing to donate organs of this patient. Ninety percent of students classified the patients in coma and in a vegetative state as alive, but still 34% of them would donate organs of those patients. The group of physicians properly determined the patients' diagnoses, but 10% of them accepted organ procurement from patients in coma and in a vegetative state. CONCLUSION: Our results supported the earlier observations of low public knowledge and inadequate understanding of brain death criteria and organ procurement processes. The majority of students were willing to accept organ procurement from severely ill but alive patients, in contrast with physicians. A considerable increase in public educational activity in this field is urgently recommended.

Superoxide anion as a marker of ischemia-reperfusion injury of the transplanted kidney.

Reactive oxygen species (ROS) are considered to be important factors involved in the pathophysiology of renal ischemia-reperfusion injury. ROS-induced alterations of proteins, carbohydrates, DNA, and lipid membranes lead to cell and organ dysfunction. Several antioxidant defense mechanisms exist to prevent or limit oxidant injury. Cellular Cu-Zn superoxide dismutase, catalase, and cellular glutathione peroxidase (cGSH-Px) are enzyme ROS scavengers, implicated in the protection against kidney damage resulting from ischemia-reperfusion injury. Reduced glutathione, a cosubstrate of cGSH-Px, have been shown to display a reductive properties without the contribution of enzymes. We examined superoxide anion (O(2)(-)) production by neutrophils, without and with stimulation using opsonized zymosan, in the whole blood of renal transplant patients before and after (5 and 15 minutes) reperfusion. The mean O(2)(-) concentration after reperfusion was statistically significantly higher than that before reperfusion.

[Evaluation of some selected structural and functional parameters of red blood cells as oxidative stress markers in elderly people]. / Ocena wybranych parametrów strukturalnych i czynnosciowych krwinek czerwonych jako markerów stresu oksydacyjnego u osób w wieku podeszlym.

UNLABELLED: The ageing process induces age-related involutionary changes and leads to increased occurrence of many diseases. One of the most important theories of ageing and development of many pathologies is the free radical theory, which assumes that ageing process leads to lost of oxidative balance. THE AIM: of the research was to evaluate the degree of membrane lipid peroxidation, internal microviscosity, activity of membrane ATPase, both total and Na(+)K(+)-dependent, and markers of oxidative damage in erythrocyte membrane protein in elderly people. MATERIAL: The examination was performed on 35 people. The examined group (15 persons, mean age 71,3) consisted of healthy elderly people. The reference group was formed with younger healthy people (20 persons, mean age 55). RESULTS: Erythrocyte membrane lipid peroxidation was found stronger in the group of elderly people. Erythrocyte internal microviscosity was significantly higher in the elderly. The activity of ATPase, both total and Na(+)K(+)-dependent, appeared remarkably greater in the group of younger people. Stronger membrane lipid damage was observed in older age group, which may be implied by lower--SH group concentration, and higher W/S parameter value. CONCLUSION: The obtained results reveal that in elderly people the intensification of oxidative stress in the entire body occurs, which may be confirmed by structural and functional oxidative erythrocyte damage. This conclusion may be significant for pathogenesis of many diseases in this period of life.

Effect of vitamin E and vitamin C supplementation on antioxidative state and renal glomerular basement membrane thickness in diabetic kidney.

The aim of this study was to analyze the effect of vitamins C and E on malondialdehyde (MDA) content and activities of key antioxidant enzymes: superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) as well as glomerular basement membrane (GBM) thickness in streptozotocin-induced diabetic kidney in rats. Wistar male rats were divided into following groups (12 rats each): the control, diabetic rats, diabetic rats whose drinking water was supplemented with vitamin C in a dose of 1.0 g/l or diet was supplemented with 200 mg of vitamin E/100 g fodder. Body weight, blood glucose and HbA1C levels and 24-hour urinary albumin excretion (UAE) were studied every week (0-12 weeks). After 6 and 12 weeks, MDA content and activities of SOD, CAT and GSH-Px were measured in the kidney homogenate supernatants. Electron micrographs of glomeruli were scanned and morphometric investigations were performed by means of computer image analysis system to compare GBM thickness. The blood glucose and HbA1C concentrations and UAE in diabetic rats were significantly higher than in the control group. An increase in the MDA level and decrease in the SOD, CAT and GSH-Px activities in the kidney of diabetic rats were observed after 6 and 12 weeks of experiment. Administration of vitamins C and E did not affect body weight, blood glucose and HbA1C levels. Both vitamin C and vitamin E decreased lipid peroxidation and augmented the activities of antioxidant enzymes studied in the kidneys of diabetic rats as well as reduced UAE, decreased kidney weight and GBM thickness. The results indicate the potential utility of antioxidant vitamins in the protection against the development of diabetic nephropathy.

The effect of verapamil on the antioxidant defence system in diabetic kidney.

BACKGROUND: The aim of this study was to examine the effect of verapamil (VP) on lipid peroxidation and activities of key antioxidant enzymes: superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px); as well as on glomerular basement membrane (GBM) thickness in streptozotocin-induced diabetic kidney in rats. METHODS: Wistar male rats were divided into three groups, 12 rats each: the control (C), diabetic rats (DR), and DR receiving VP, 7 mg/kg body weight in drinking water (DR + VP). Blood glucose (BG) and HbA(1c) levels, 24-h urinary albumin excretion (UAE) and body weight (BW) were measured every week (0-12 weeks). After 6 and 12 weeks, the animals were sacrificed and malondialdehyde (MDA) content and activities of SOD, CAT and GSH-Px were determined in the kidney homogenate supernatants. Electron micrographs of the glomeruli were scanned and morphometric investigations were performed by means of a computer image analysis system to compare the glomerular basement basal membrane (GBM) thickness. RESULTS: The levels of BG, HbA(1c) and UAE in DR were significantly higher than in the C group. A progressive increase in the MDA level and a decrease in the SOD, CAT and GSH-Px activities in the kidney of DR were observed after 6 and 12 weeks. VP administration did not affect BW changes, BG and HbA(1c) levels in DR. VP decreased lipid peroxidation and augmented the activities of antioxidant enzymes studied in the kidneys of DR as well as decreased kidney weight, GBM thickness and albuminuria in DR. CONCLUSIONS: These results confirm the role of oxidative stress in the development of diabetic nephropathy and point to the possible antioxidative mechanism of the nephroprotective action of VP.

[Evaluation of oxygen free radical generation and antioxidative enzymatic activity in blood of patients with carcinoma of the larynx (preliminary report)]. / Ocena generacji anionorodnika ponadtlenkowego oraz antyoksydacyjnej aktywnosci enzymatycznej we krwi obwodowej chorych z rakiem krtani (doniesienie wstepne).

The aim of the study were the evaluation of generation of oxygen free radical and antioxidative enzymatic activity in blood of patients with carcinoma of larynx (7 persons, aged 48-68) in comparison with healthy persons (15, aged 21-28). The superoxide anion generation of blood granulocytes described by the rate of the reducted cytochrom C at rest and after stimulation with opsonized zymosan according to Bellavite et al. were measured. The enzymatic activity of superoxide dismutase in blood (method of Misra and Fridovich), catalase (method of Beers and Sizer) and malonyl dialdehyde (method of Placer et al.). The obtained data pointed at the growth of generation of oxygen free radical in blood (at rest and after stimulation with opsonized zymosan) of patients with carcinoma of larynx in comparison with the healthy ones. At the same patients with carcinoma of larynx observed at significant reduction of antioxidative enzymatic activity against the oxygen species generation in blood and significant growth of peroxidation lipids and disturbances of function of cells the human body.

[Estimation of plasma malonyl dialdehyde concentration in patients with pulmonary tuberculosis]. / Ocena stezenia dialdehydu malonowego w osoczu chorych na gruzlice pluc.

Plasma malonyl dialdehyde (MDA) concentration was determined by Placer method in patients with pulmonary tuberculosis before and during treatment period with tuberculostatic drugs. Patients were divided into three groups. Group I comprised 20 patients (10 men and 10 women) aged 24-74 years (mean 48.3 years) with pulmonary tuberculosis BK(+). Group II comprised 20 patients (15 men and 5 women) aged 19-67 years (mean 45.1 years) with pulmonary tuberculosis BK(0). The control group consisted of 16 clinically healthy persons (12 men and 4 women) aged 28-59 years (mean 42.5 years). In patients blood samples (5 ml) were collected for examination from cubital vein before, after 1-month and 2-month treatment period with tuberculostatic drugs. In the control group blood samples were collected from cubital vein once. Results of our study showed that plasma MDA concentrations in patients with pulmonary tuberculosis, both before and during treatment period with tuberculostatic drugs, were significantly higher, in comparison to the control group.

[Antioxidant enzymatic activity in blood of patients with chronic hypertrophy of the maxillary sinuses and larynx and the carcinoma of the larynx]. / Antyoksydacyjna aktywnosc enzymatyczna we krwi obwodowej chorych z przewleklym przerostowym zapaleniem zatok szczekowych i krtani oraz rakiem krtani.

The aim of the work was the examinations of antioxidant enzymatic activity in blood of patients with chronic hypertrophic of maxillary sinusitis (15 patients) and laryngitis (10 patients), and carcinoma of larynx (7 patients) in comparison with healthy persons (15). The enzymatic activity of superoxide dismutase in blood (method of Misra and Fridovich), katalase (method of Beers and Sizer) and malonyl dialdehyde (method of Placer) were evaluated. The obtained results pointed at significant reduction of antioxidant enzymatic activity against the oxygen species generation in blood of patients with chronic hypertrophic of maxillary sinusits and laryngitis and mainly with the carcinoma of larynx (in comparison with healthy persons). At the same patients observed the significant growth of peroxydation lipoid and disturbances of function of cells the human body.

Changes in the structure and properties of corpus vitreous and the preservation of enzymatic composition of rabbit antioxidative system under the influence of sulphur hexafluoride gas.

PURPOSE: To evaluate clinical, physiochemical and biochemical changes in rabbit vitreous body caused by local injection of sulphur hexafluoride gas. MATERIAL AND METHODS: The volume of fluid vitreous fraction was measured with Na+, K+, Mg2+, Ca2+ levels and full proteins concentration in both vitreous fractions in 24 New Zealand rabbits at 2, 7 and 14 day after SF6 injection. The activity of superoxide dismutase, catalase and malonyl dialdehyde were used to evaluate the activity of antioxidative enzymatic system. Control group consisting of 6 New Zealand rabbits had no experimental procedures. RESULTS: In the investigated group, the fluid vitreous fraction volume was increased while gelatous one was diminished from 0.08 ml in control group to 0.32 ml in the study group (on day 14). The level of Na+, K+, Mg2+, Ca2+ in the fluid fraction was unchanged. On day 7, we noticed statistically significant increase in protein concentration in comparison with the control group and the study group on 14 day. The activity of superoxide dismutase and catalase as well as the level of malonyl dialdehyde were increased in the fluid vitreous fraction compared to the gelatous one in the control group. After the SF6 injection we did not observe any changes of superoxide dismutase and catalase activities in the gelatous part of vitreous body while in the fluid one there was statistically significant decrease in the enzymatic activity and the MDA level in the whole observation time. CONCLUSIONS: The injected sulphur hexafluoride gas caused the damage of the gelatous vitreous fraction with the increase in the fluid one. The oxygen free radicals might trigger these pathological processes.

Moderate systemic hypothermia and cold crystalloid cardioplegia influence on myocardial ischemic and revascularisative injury.

The number of granulocytes, their ability to generate superoxide anion (O2-) and the activities of Cu, Zn--superoxide dismutase (SOD-1), glutathione peroxidase (GSH-Px), catalase (CAT) as well as malonyldialdehyde (MDA) concentrations in erythrocytes in the blood extracted from the venous sinus and aorta under coronary artery bypass were examined with the use of St. Thomas Hospital cardioplegic solution. Specimens at the peak of ischemia of the right atrium for ultrastructural examination of the endothelial cells of capillary vessels and sarcomers were taken. The blood was obtained during cardiopulmonary bypass (CPB) before the aorta clamping and immediately after aorta declamping (peak of ischaemia) between 1-3 minute and 10-13 minute of reperfusion. Increase of the number of granulocytes both in the coronary sinus and aortal blood at all examined intervals as well as decrease in the number of ones in sinus compared with aortal blood was noted. The ability to produce superoxide anion radical decreased at the peak of ischemia and during reperfusion. The activity of SOD-1 was lower both after the period of ischemia and reperfusion. The increase in aortal blood activity during reperfusion was characteristic of GSH-Px; the activity was higher in the blood sample from the coronary sinus taken during ischemia and initial reperfusion. With the exception of the initial reperfusion the activity of CAT diminished in all observed cases. MDA concentration did not demonstrate any significant changes with the exception of the initial reperfusion in the aortal blood and later towards the end of reperfusion in the blood from the coronary sinus. Ultrastructural studies indicated overhydration of the cells both in the endothelium and the intercellular space. The obtained data demonstrate that the applied cardioplegic solution protects the myocardium from harmful effects of reactive oxygen species produced as a result of ischemia and reperfusion.

[Assessment of nitric oxide level in aqueous humor under physiological conditions and after lens extraction with PMMA during experimental work in rabbits]. / Ocena stezenia tlenku azotu w cieczy wodnistej w warunkach fizjologicznych oraz po usunieciu soczewki i wszczepie sztucznej soczewki z PMMA w badaniach doswiadczalnych u królika.

PURPOSE: To evaluate the presence of nitric oxide and measure its level in the aqueous humor of the rabbit's eye, in physiological conditions and after extracapsular lens extraction and PMMA artificial lens implantation. We also investigated nitric oxide maintenance during early postoperative period (between 1-5 day after surgery). MATERIAL AND METHODS: We used 30 rabbits (weighing 3.0-3.5 kg) Just before surgery samples of aqueous humor were aspirated by anterior chamber puncture. Lens was extracted with extracapsular (envelope) technique. In 15 eyes PMMA IOL was implanted in the bag and 15 eyes were left aphakic. The aqueous samples were collected on 1st, 3rd, 5th days after surgery. Nitric oxide in each sample was measured with respect to fluorometric assay. RESULTS: In aqueous humor in physiological conditions we detected nitric oxide. Its level was estimated on the value of 26.52 nM/dl. After extracapsular lens extraction in both groups the level of nitric oxide was significantly higher than in control group. The day and value of NO level was different among examined groups. Nitric oxide level diminished significantly on 5th postoperative day. CONCLUSION: We came to conclusion that after ECCE and PMMA IOL implantation NO level was significantly higher as compared with control. This higher NO level after lens extraction can be responsible for the blood aqueous breakdown.

[Assessment of nitric oxide level in aqueous humor under physiological conditions and after lens extraction and artificial foldable acrylic lens implantation during experimental work in rabbits]. / Ocena stezenia tlenku azotu w cieczy wodnistej w warunkach fizjologicznych oraz po usunieciu soczewki i wszczepie sztucznej zwijalnej soczewki akrylowej w badaniach doswiadczalnych u królika.

PURPOSE: To evaluate the presence of nitric oxide (NO) and its level in the aqueous humor of rabbit's eye physiological conditions and after phacoemulsification and acrylic foldable artificial lens implantation. We also investigated nitric oxide maintenance during early postoperative period (between 1-5 days after surgery). MATERIAL AND METHODS: We examined 30 rabbits (weighing 3.0-3.5 kg) Just before surgery samples of aqueous humor were aspirated. Lens was extracted with phacoemulsification technique. In 15 eyes acrylic foldable IOL (group III) was implanted and 15 eyes were left aphakic (group IV). The aqueous samples were collected on 1, 3, 5 days after surgery. Nitric oxide in each sample was determined with fluorometric assay. RESULTS: The level of NO in aqueous humor in physiological conditions was estimated to 26.52 nM/dl. After phacoemulsification in both groups the level of nitric oxide was higher than in control group. The day and value of the highest NO level was different among examined groups. The highest level of NO was released during 1st day in group III and on the 3rd day in the group IV. CONCLUSION: We came to conclusion that the level of nitric oxide in aqueous humor after phacoemulsification in both groups is higher than in control group but significantly lower than in previously examined groups in which the surgery was made with extracapsular technique.

[Changes in the antioxidant system of the vitreous in rabbits after administration of sulfur hexafluoride]. / Zmiany w ukladzie antyoksydacyjnym ciala szklistego królików po podaniu szesciofluorku siarki.

PURPOSE: To evaluate the enzymatic activity of antioxidant system of rabbit's vitreous after sulfur hexafluoride (SF6) application. MATERIAL AND METHODS: Activity of CuZn-SOD, catalase and concentration of MDA in fluid and gel fraction of vitreous were determined in 24 rabbits of New Zealand race on the 2nd, 7th and 14th day after SF6 application. Control group consisted of 6 animals which did not undergo any operations. RESULTS: Dismutase and catalase activity as well as MDA concentration were higher in fluid fraction than in gel fraction in animals of control group. After SF6 application the activity of enzymes and MDA concentration did not change, whereas in fluid fraction all these values were statistically significantly reduced in all time intervals. CONCLUSIONS: SF6 leads to disintegration of vitreous structure especially just after its application. Damage to hyalocytes causes dysfunction of enzymatic system. Specific fluid fraction structure and insufficient number of substrates for peroxidation processes are the reasons for simultaneous reduction of MDA concentration.

[Changes in the antioxidant system of the aqueous humor, lens and erythrocytes after sulfur hexafluoride application to the vitreous of rabbits]. / Zmiany w ukladzie antyoksydacyjnym cieczy wodnistej, soczewce i krwinkach czerwonych krwi obwodowej po podaniu szesciofluorku siarki do ciala szklistego królików.

AIM: To determine the activity of superoxide dismutase, catalase and the concentration of malondialdehyde (MDA) in aqueous humor, lens and red blood cells after application of sulfur hexafluoride (SF6) into vitreous of rabbits. MATERIAL AND METHODS: 0.5 ml of 100% SF6 was injected into the vitreous of 24 rabbits of New Zealand race. The animals were randomly divided into 3 groups (of 8 rabbits each) depending on the observation day: group 1-2nd day of experiment, group 2-7th day and group 3-14th observation day. The control group (gr. 0) consisted of 6 rabbits that did not undergo any operations. Activity of superoxide dismutase, catalase and MDA concentration were determined in aqueous humor, lens and systemic blood erythrocytes. RESULTS: On the 7th day of observation an increased activity of dismutase and catalase as well as simultaneous increased MDA concentration were observed. In the lens on the 7th day the increased activity of dismutase was significant in relation to the results in the next time interval, whereas MDA concentration was significantly lower in all time intervals of the experiment in comparison with control group. In erythrocytes an increased activity of catalase was noticed on the 2nd and 14th day. CONCLUSIONS: Increased occurrence of active oxygen species in aqueous humor leads to insufficiency of the antioxidant system and intensification of peroxidation processes, which is reflected by increased MDA concentration. However, in the lens of this experimental model a slight stimulation of antioxidant system by a small number of free radicals is observed, which provokes a reaction of sweeping them away. Efficiency of lens antioxidant system is secured by weakening of peroxidation processes, which is expressed in minimal drop of MDA concentration.

Luminol luminescence induced by oxidants in antioxidant-deficient yeasts Saccharomyces cerevisiae.

Luminol chemiluminescence induced in the presence of yeast cells and yeast cell homogenates was significantly induced by exogenous oxidants (hydrogen peroxide and menadione). tert-Butyl hydroperoxide did not stimulate chemiluminescence by itself but augmented menadione-induced chemiluminescence. Comparison of yeast strains deficient in catalase, superoxide dismutase or glutathione showed that only glutathione-deficient strains showed elevated chemiluminescence in this system. These results support the idea that more reactive species than hydrogen peroxide and superoxide are critical in the induction of luminol chemiluminescence.

Effect of the H2 histamine receptor antagonist on oxygen metabolism in some morphotic blood elements in patients with ulcer disease.

BACKGROUND/AIMS: Our investigations was carried out in order to examine the effect of cimetidine, ranitidine and famotidine on the generation of free radicals, lipid peroxidation and enzymatic antioxidative defense in the blood of patients with peptic ulcer disease, clinically diagnosed as gastric or duodenal ulcer. METHODOLOGY: 124 non-smoking males (aged 20-51 years), were randomly divided into 4 groups: 28 patients received intravenously 200 mg of cimetidine; 26 patients intravenously 50 mg comprised of ranitidine; 25 patients received intravenously 20 mg of famotidine; and 45 healthy men served as the control group. Superoxide dismutase activity, malonyldialdehyde concentration in blood platelets and superoxide anion generation in granulocytes were determined in all examined men. An assay of superoxide dismutase activity and malonyldialdehyde concentration were performed before drug administration and after 2 and 72 hours. Superoxide anion generation was estimated before drug administration and after 2 hours. RESULTS: Our data indicate that all examined H2 receptor antagonists stimulate superoxide dismutase activity, but after 72 hours a distinct increase was observed, in addition to a decrease of malonyldialdehyde concentration. No differences have been observed in superoxide anion generation in patients with ulcer disease or in healthy subjects before and after ranitidine and famotidine administration. Only after 2 hours of cimetidine administration was a significant increase in superoxide anion generation observed. CONCLUSION: We concluded that H2 receptor antagonists have a beneficial effect on antioxidative processes.

Free radicals generation by granulocytes from men during bed rest.

Phagocytes (granulocytes and macrophages) are one of the most important sources of reactive oxygen species (ROS) in the human body. A stimulated phagocyte reaches within a dozen of seconds the functional state known as the respiratory burst. This phenomenon includes three interrelated processes: 1) a several fold increase in the oxygen uptake, 2) generation of large amounts of ROS, and 3) stimulations of glucose metabolism via the pentose phosphate shunt. The main ROS generated during phagocytosis is the superoxide radical anion O2-. The reaction of dismutation of O2- generates hydrogen peroxide (H2O2). O2- and H2O2 do not have strong cytocidal proerties but are supbstrates for the generation of more active biologically ROS, especially the hydroxyl radical (OH) and hypochlorous acid (HOCl). A stimulated phagocyte (especially neutrophil) generates also vast amounts of nitric oxide (NO-). NO- may react with the superoxide radical anion forming peroxynitrate (ONOO-). Reactivity of ONOO- is comparable to that of the hydroxyl radical. ONOO- plays probably a very important role in the so-called oxygen-dependent killing mechanisms during phagocytosis. The role of NO- in the process of phagocytosis is unclear. However, NO-, apart from other biological functions, inhibits aggregation and adhesion of blood platelets. Morevover, it seems to participate in the slefregulation of granulocyte cativity (probably limiting excessive inflammative reaction). Excess of ROS generated by phagocytes may damage biologically important macromolecules. Membrane lipids are particularly vulnerable to peroxidation. It leads to alterations in membrane fluidity and permeability. ROS excreted to the environment of a phagocyte may impair functions of other morphotic elements of blood (erythrocytes and thromobcytes) in this way. Bed rest covers two conditions: 1) limitation of motoric activity (hypodynamia) and 2) restraining the pressure on bones along parallelly to their long axes (hypogravia). Under bed rest conditions organism is subject to many harmful changes. I.a., bed rest affects also leukocyte functions. However, we were not aware of any sutdies concerning the effect of bed rest on the respiratory burst of granulocytes. This study was aimed at an examination of the effect of bed rest on the generation of O; and H2O2 by neutrophils (PMNL).

Effect of long term bed rest in men on enzymatic antioxidative defence and lipid peroxidation in erythrocytes.

Bed rest is an integral part of treatment of numerous diseases. Typical examples are bone fractures of lower extremities and pelvis. Temporary immobilization is necessary also, e.g., in heart diseases (stroke), backbone and imminent abortion. The sick organism spares energy during the bed rest wich is beneficial. However, bed rest results in many alterations which are disadavantageous. They concern the function of almost all organs and systems but affect most significantly the locomotor and ciruclatory systems. Bed rest brings also about changes in the composition of peripheral blood and functions of the morphotic elements of blood. Red blood cells are subjected to the action of large amounts of reactive oxygen species (ROS). During oxidation of hemoglobin to methemoglobin superoxide radical anion (O2-) is formed: HbFe2+ + O2 --> MetHbFe3+ + O2- (1) Ferrous and ferric ions present in the cytoplasm of red blood cells may be catalysts of the Fenton reaction leading to the production of the hydroxyl radical: O2- + Fe3+ --> O2- + Fe2+ (2) Fe2+ + H2O2 --> Fe3+ + OH + HO- (3) OH shows a tremendous reactivity. It may react with lipids, proteins, nucleic acids and carbohydrates. The process of lipid peroxidation is best understood. It concerns mainly polyunsaturated fatty acids present in cell membranes. Peroxidation of membrane lipids decreases membrane fluidity and impairs its barrier function. The lowered membrane fluidity compromises erythrocyte deormability which in turn disturbs oxygen delivery to the tissues. End productions of lipid peroxidation are low-molecular wieght compounds, among them carbohydrates (ethane and pentane) and aldehydes, e.g. malondialdehyde (MDA). MDA concentration is an acknowldeged marker of the intensity of lipid peroxidation. Erythrocytes contain a complex system of protection against the action of ROS. It includes various enzymatic and non-enzymatic mechanism. The most important antioxidative enzymes of the red blood cells are superoxide dismutase (Cu,Zn-SOD, EC 1.15.1.1) catalase (CAT, EC 1.11.1.6) and glutathione peroxidase (GSH-Px, EC 1.11.1.9). Cu,Zn-SOD catalyzes the dismuation of O2- to hydrogen peroxide (H2O2). Catalase and peroxidase remove H2O2 and, moreover, GSH-Px can reduce lipid peroxides. Under normal conditions an equilibrium exists between the formation and removal ROS. If ROS are formed in excess or the defensive antioxidative mechanism are inefficient, oxidative stress develops. Derangement of the equilibrium between the formation and removal of ROS is important in the pathosgenesis of many diseases, e.g. atherosclerosis, diabetes, Down syndrome and Alzheimer disease. There are literature data on disturbances of enzymatic antioxidant defense mechanism of blood plateless during bed rest. This study was aimed at an examination of the post-traumatic bed rest on the enzymatic antioxidative defense mechanisms and lipid peroxidation in erythrocytes.