Efekat MnSOD (E. coli) Na relaksaciju izolovane renalne arterije pacova izazvanu natrijum-nitro-prusidom

In this study themolecular foundation of nitric oxide induced relaxation of arteries, with or without endothelium, of normotensive and spontanously hypertensive ratswas re-examined. With this purpose in mind, the effects of the nitric oxide donor sodium nitroprusside (NaNP), with and without manganese containing superoxide dismutase (MnSOD E.C. 1.15.1.1), on rat renal artery relaxation was strudied. The results show that the relaxation effect of NaNP is two times higher in normotensive, compared to spontaneously hypertensive rats. Similar differences exist in the relaxation effects of NaNP on isolated renal arteries without endothelium, indicating that besides the difference in the function of an endothelium, concerning basal NO production in normotensive and hypertensive rats, there is a differencewith respect to NO relaxation in the smoothmuscle that is induced by hypertension. MnSOD decreased the relaxation effect of NaNP in all the examined renal arteries, more in normotensive than in hypertensive ones regardless of the presence of an endothelium. These results show that MnSOD, by modifying the chemical versatility of NO into redox active forms - nitrosonium (NO+) and nitroxyl (NO-), produces different relaxation effects in normotensive and hypertensive arteries of rats, with or without an endothelium, potentiating the role of nitroxyl induced relaxation in sponteneously hypertensive rats. The results prove the need for the synthesis of complex NO donors, as the mechanisms of artery relaxation are different due to an endothel and smooth mouscle changes in hypertensive, as compared to normotensive rats.U ovom radu smo pokušali da detaljnije ispitamo molekulsku osnovu azotoksid indukovane relaksacije arterija, sa i bez endotela, normotenzivnih i spontano hipertenzivnih pacova. U tu svrhu ispitivan je efekat azot-oksid (NO) donora natrijum- nitroprusida (NaNP), bez i u prisustvu superoksid-dismutaze koja sadrži mangan (MnSOD, EC 1.15.1.1) na relaksaciju renalne arterije. Rezulati pokazuju dvostruko veći relaksantni efekat NaNP kod normotenzivnih, u odnosu na spontano hipertenzivne pacove. Slična razlika postoji i u relaksantnom efektu NaNP na izolovane renalne arterije normotenzivnog i hipertenzivnog pacova kada je odstranjen endotel što ukazuje da, pored razlike u funciji endotela u odnosu na bazalnu NO produkciju kod normotenzivnih i spontano hipertenzivnih pacova, postoje i promene u glatkim mišićima indukovane hipertenzijom, u odnosu na NO relaksaciju. MnSOD kod svih grupa smanjuje relaksantni efekat SNP i to više kod normotenzivnih nego kod hipertenzivnih nezavisno od prisustva endotela. Ovi rezultati pokazuju da MnSOD menjajući hemijsku prirodu iz NaNP oslobođenog NO u redoks aktivne forme – nitrozonijum (NO+) i nitroksil (NO–) – ostvaruje različit relaksantni efekat kod normotenzivnih i kod hiperetenzivnih pacova sa i bez endotela potencirajući značaj nitroksilom indukovane relaksacije kod spontano-hipertenzivnih pacova. Dobijeni rezultati potvrđuju potrebu sinteze NO donora koji daju različite redoks-aktivne forme NO budući da se primarni mehanismi relaksacije razlikuju u zavisnosti od karakteristika endotela i glatkih mišića, kod hipertenzije u odnosu na normotenzivno stanje

Elevated serum uric acid reduce heart damage in patients undergoing open-heart surgery

Objectives: Peroxynitrite is species claimed to propagate ischemia/reperfusion damage. In this report levels of serum uric acid (UA), a peroxynitrite scavenger, are compared with creatine phosphokinase (CPK) in male patients before and after open-heart surgery in order to asses if increased levels of UA may protect heart from biochemical damage induced by peroxynitrite during the coronary by-pass grafting (CABG) intervention. Methods: 45 male patients (16 carvedilol pretreated (6.25 mg/ daily, during 6 weeks before surgery, mean age 55.3±1.7 years, range 50-71) and 29 patients without carvedilol pretreatment (mean age 58.3±1.4 years, range 47-73) underwent elective CABG were examined. Study inclusion criteria were CABG performed on two and more coronary-vessels with aortic crossclamp during 30-40 minutes. For assessment of patients objective health status before operations Euro- SCORE were used. Serum uric acid (UA) levels and creatine phosphokinase (CPK) were measured spectrophotometricaly by using a quantitative enzymatic assay. Results: Carvedilol pretreated patients had higher amount of serum UA (p<0.05) comparing to non-treated patients. During the surgical procedure patients are subjected to temporary ischemia due to transfer from corporeal to extracorporeal circulation. In this period of time the amount of UA decreased in carvedilol pretreated group (406±46 (t1) vs. 300±22 mmol/L (t2)) to the level of non-treated patients (328±14 (t1) vs. 322±18 mmol/L. Carvedilol pretreated patients and non-treated patients had the same level of CPK at the beginning of the surgical procedure (t1) (78±6 vs. 83+13 U/L) but lower increase (p<0.05) in CPK activity in carvedilol pretreated patients in respect to nontreated patients (338±46 vs. 644±103 U/L) at the end of procedure (t2). Such results suggest that open heart surgery led to elevated CPK levels, but this effect was less pronounced in patients with higher level of UA. Conclusions: Our results suggest possible role of UA in the protection from reperfusion injury. Increase of UA before surgery may be beneficial factor during CABG procedure in patients treated with carvedilol by decreasing level of peroxynitrite as one of molecular causes of reperfusion injury. Our results showed influence of UA on CPK levels at the end of surgical procedure, indicating that increased levels of UA may protect heart from biochemical damage induced by peroxynitrite during the CABG intervention.Ciljevi: Postoji mišljenje da Peroxynitrite širi - pojačava oštećenja koja potiču od ishemije /reperfuzije. U ovom radu poređen je nivo mokraćne kiseline (UA) u serumu, sakupljača peroksinitrita, sa nivoom kreatin fosfokinaze (CPK) kod muškaraca pre i nakon operacije na otvorenom srcu, a u cilju procene da li povećani nivoi UA mogu služiti kao zaštita od biohemijskih oštećenja izazvanih upotrebom peroksinitrita tokom hirurške CABG intervencije. Metode: Ispitivano je 45 bolesnika (16 je prethodno primalo karvedilol - 6.25 mg dnevno, tokom 6 nedelja pre operacije, prosečne starosti 55.3±1.7 godina, 55-71, i 29 bolesnika koji nisu primali karvedilol, prosečne starosti 58.3±1.4 godina, 47-73) koji su bili podvrgnuti CABG hirurškoj intervenciji. Kriterijumi za uključenje u studiju su bili izvođenje CABG na dva ili više sudova sa klemovanjem aorte u trajanju od 30 - 40 minuta. Za procenu zdravstvenog stanja bolesnika pre operacije korišćen je EuroSCORE. Nivoi UA i kreatin fosfokinaze (CPK) u serumu su mereni uz pomoć spektrofotometrije korišćenjem kvantitativnog enzimskog eseja. Rezultati: U bolesnika koji su prethodno primali karvedilol uočeni su uvećani nivoi UA u serumu (p< 0.05) u poređenju sa bolesnicima koji nisu primali pomenuti lek. Tokom hirurške procedure bolesnici su podvrgnuti privremenoj ishemiji zbog prebacivanja sa telesnog na vantelesni krvotok. U tom periodu vrednosti UA su snižene u bolesnika koji su prethodno primali karvedilol (406+46 (t1) naspram 300±22 mol/L (t2)) bolesnika koji ga nisu primali (328±14 (t1) naspram 322±18 mol/L). Bolesnici koji su primali karvedilol i oni koji nisu imali su isti nivo CPK na početku hirurške procedure (t1) (78±6 naspram 83±13 U/L) ali je na kraju procedure uočen niži porast aktivnosti CPK (p<0.05) u bolesnika koji su primali karvedilol u poređenju sa bolesnicima koji nisu (338±46 prema 644±103 U/L) (t2). Ovakvi rezultati sugerišu da intervencija na otvorenom srcu dovodi do povećanja nivoa CPK, ali je taj efekat bio manje izražen u bolesnika koji su imali viši nivo UA. Zaključak: Naši rezultati ukazuju na moguću pozitivnu ulogu mokraćne kiseline u zaštiti od oštećenja koja može prouzrokovati reperfuzija. Povećanje nivoa UA pre operacije može predstavljati koristan faktor tokom CABG procedure u bolesnika koji su primali karvedilol, jer snižava nivo peroksinitrita kao jedan od uzroka oštećenja od reperfuzije. Naši rezultati pokazuju uticaj UA na nivoe CPK pri kraju hirurške intervencije, što pak ukazuje na to da uvećani nivoi UA mogu štititi srce od biohemijskih oštećenja izazvana peroksinitritom tokom CABG intervencije

Effect of bovine CuZn superoxide dismutase on C3 clone of B-16 mouse melanome cells in culture

B-I6/C3 mouse melanoma cells undergo melanogenesis and differentiation 9 days after plating under usual conditions. In our experiments the effect of exogenous bovine CuZn superoxide dismutase (CuZn SOD) on B-I6/C3 cells in cultures was studied. The exogenous CuZn SOD was added, 24 hours after cell plating, in growth medium, which either contain or not Fetal Calf Serum (FCS). B-I6/C3 melanoma cells growth was followed over 5 days. Different effect of CuZn SOD on the culture was observed in relation to FCS present in growth medium. CuZn SOD induced a dose dependent increase in melanin content of B-I6/C3 cells in serum deprived medium. Our results are discussed in respect to dismutating CuZn SOD activity, which may act to an enhanced level of oxidative stress, related to the higher metabolism in melanoma cells. On the other hand we supposed that CuZn SOD produced an elevated level of hydrogen peroxide. Therefore hydrogen peroxide than may play some role in cell differentiation as transmembrane messenger.Ćelije B-I6/C3 melanoma miša melaniziraju i diferenciraju devetog dana posle zasađivanja pod uobičajenim uslovima. U našim eksperimentima smo proučavali uticaj egzogeno dodavane CuZn superoksid-dismutaze ( CuZn SOD) na ćelije B-I6/C3 u kulturi. Egzogena CuZn SOD je dodavana 24 časa posle zasađivanja ćelija u medijume za rast koji su sadržavali ili ne goveđi fetalni serum. Rast ćelija B-I6/C3 je praćen pet dana. Zapažen je različit uticaj CuZn SOD na ćelijske kulture gajene sa serumom ili bez njega. CuZn SOD izaziva dozno zavisnu sintezu i nakupljanje melanina u ćelijama koje su rasle u medijumu koji nije sadržavao goveđi fetalni serum. Naši rezultatu se mogu diskutovati u svetlu dismutacione aktivnosti CuZn SOD koja deluje na povećani nivo oksidacionog stresa izazvanog visokim metabolizmom melanoma ćelija. S druge strane, pretpostavili smo da CuZn SOD svojom aktivnošću proizvodi više vodonik peroksida. Vodonik peroksid, nadalje, može da ima izvesnu ulogu u diferencijaciji ćelija kao transmembranski signalni molekul.Projekat ministarstva br. 166

Activity of antioxidative defense enzymes in the blood of patients with liver echinococcosis

Background/Aim. Chronic echinococcocal disease is the parasite human disease caused by the penetration of larval (asexual) stages of the canine tapeworm (Echinococcus granulosus) in the liver of humans. After the penetration of the parasite, the host organism react by activating complement- depending immune response. The aim of this study was to elucidate the influence of larval form of Echinococcus granulosus in the liver on the activity of antioxidative defense enzymes in the blood of patients before and after the surgical intervention. Methods. We investigated the activity of antioxidative defense enzymes: copper/zinc containing superoxide dismutase (CuZn SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione reductase (GR) and glutathione-S-transferase (GST) in the blood of patients before and after the surgical intervention in respect to the controls, clinically healthy persons. Results. Our results showed that the activity of the GSH-Px was significantly decreased in the plasma of the patients with echinocococal disease before the surgery in respect to the controls. The activity of GST was significantly higher in the blood of the patients after the surgery in comparison to the controls. Conclusion. Chronic liver echinoccocal disease caused significant changes of some antioxidative defense enzymes, first of all Se-dependent enzyme GSH-Px, which could be a suitabile biomarker in the biochemical evaluation of the disease. This work represents a first comprehensive study of the activity of antioxidative defense enzymes in cronic liver echinococcocosis in the patients before and after the surgical intervention in respect to the clinically healthy persons

Glutathione status in the blood of rats after reticulocytosis induced by phenylhydrazine and bleeding

In this experiment, we compared the in vivo effects of phenylhydrazine (PHZ) and bleeding treatment on the redox status and glutathione antioxidative mechanism parameters in the plasma and red blood cells (RBC) of rats. Results showed a lower level of reactive oxygen species (ROS), a higher level of lipid peroxidation and the effective antioxidative role of the glutathione system in the blood of bleeding rats. PHZ-treatment induced higher concentrations of ROS and an accumulation of oxidized glutathione in the plasma, while the glutathione system showed a satisfactory antioxidative capacity in the RBC of rats. When comparing the two anemic groups, the PHZ-treated rats showed marked oxidative stress in the plasma.Cilj ovog rada je bio da se isprate in vivo efekti fenilhidrazinom (PHZ) i krvarenjem indukovane retikulocitoze na parametre redoks i glutationskog antioksidativnog statusa u plazmi i crvenim krvnim ćelijama (RBC) pacova. Rezultati pokazuju niži nivo reaktivnih vrsta kiseonika (ROS), viši nivo lipidne peroksidacije i efikasnu antioksidacionu ulogu glutationskog sistema u krvi iskvavljenih pacova. Tretman PHZom prouzrokovao je više koncentracije ROSa i akumulaciju oksidovanog glutationa u plazmi, dok je glutationski sistem pokazao efikasan antioksidativni kapacitet u RBCu pacova. Kada se uporede dve anemične grupe, izraženiji oksidacioni stres se javlja u plazmi pacova tretiranih PHZom.Projekat ministarstva br. 143035

Elevated serum uric acid reduce heart damage in patients undergoing open-heart surgery

Objectives: Peroxynitrite is species claimed to propagate ischemia/reperfusion damage. In this report levels of serum uric acid (UA), a peroxynitrite scavenger, are compared with creatine phosphokinase (CPK) in male patients before and after open-heart surgery in order to asses if increased levels of UA may protect heart from biochemical damage induced by peroxynitrite during the coronary by-pass grafting (CABG) intervention. Methods: 45 male patients (16 carvedilol pretreated (6.25 mg/ daily, during 6 weeks before surgery, mean age 55.3±1.7 years, range 50-71) and 29 patients without carvedilol pretreatment (mean age 58.3±1.4 years, range 47-73) underwent elective CABG were examined. Study inclusion criteria were CABG performed on two and more coronary-vessels with aortic crossclamp during 30-40 minutes. For assessment of patients objective health status before operations Euro- SCORE were used. Serum uric acid (UA) levels and creatine phosphokinase (CPK) were measured spectrophotometricaly by using a quantitative enzymatic assay. Results: Carvedilol pretreated patients had higher amount of serum UA (p<0.05) comparing to non-treated patients. During the surgical procedure patients are subjected to temporary ischemia due to transfer from corporeal to extracorporeal circulation. In this period of time the amount of UA decreased in carvedilol pretreated group (406±46 (t1) vs. 300±22 mmol/L (t2)) to the level of non-treated patients (328±14 (t1) vs. 322±18 mmol/L. Carvedilol pretreated patients and non-treated patients had the same level of CPK at the beginning of the surgical procedure (t1) (78±6 vs. 83+13 U/L) but lower increase (p<0.05) in CPK activity in carvedilol pretreated patients in respect to nontreated patients (338±46 vs. 644±103 U/L) at the end of procedure (t2). Such results suggest that open heart surgery led to elevated CPK levels, but this effect was less pronounced in patients with higher level of UA. Conclusions: Our results suggest possible role of UA in the protection from reperfusion injury. Increase of UA before surgery may be beneficial factor during CABG procedure in patients treated with carvedilol by decreasing level of peroxynitrite as one of molecular causes of reperfusion injury. Our results showed influence of UA on CPK levels at the end of surgical procedure, indicating that increased levels of UA may protect heart from biochemical damage induced by peroxynitrite during the CABG intervention.Ciljevi: Postoji mišljenje da Peroxynitrite širi - pojačava oštećenja koja potiču od ishemije /reperfuzije. U ovom radu poređen je nivo mokraćne kiseline (UA) u serumu, sakupljača peroksinitrita, sa nivoom kreatin fosfokinaze (CPK) kod muškaraca pre i nakon operacije na otvorenom srcu, a u cilju procene da li povećani nivoi UA mogu služiti kao zaštita od biohemijskih oštećenja izazvanih upotrebom peroksinitrita tokom hirurške CABG intervencije. Metode: Ispitivano je 45 bolesnika (16 je prethodno primalo karvedilol - 6.25 mg dnevno, tokom 6 nedelja pre operacije, prosečne starosti 55.3±1.7 godina, 55-71, i 29 bolesnika koji nisu primali karvedilol, prosečne starosti 58.3±1.4 godina, 47-73) koji su bili podvrgnuti CABG hirurškoj intervenciji. Kriterijumi za uključenje u studiju su bili izvođenje CABG na dva ili više sudova sa klemovanjem aorte u trajanju od 30 - 40 minuta. Za procenu zdravstvenog stanja bolesnika pre operacije korišćen je EuroSCORE. Nivoi UA i kreatin fosfokinaze (CPK) u serumu su mereni uz pomoć spektrofotometrije korišćenjem kvantitativnog enzimskog eseja. Rezultati: U bolesnika koji su prethodno primali karvedilol uočeni su uvećani nivoi UA u serumu (p< 0.05) u poređenju sa bolesnicima koji nisu primali pomenuti lek. Tokom hirurške procedure bolesnici su podvrgnuti privremenoj ishemiji zbog prebacivanja sa telesnog na vantelesni krvotok. U tom periodu vrednosti UA su snižene u bolesnika koji su prethodno primali karvedilol (406+46 (t1) naspram 300±22 mol/L (t2)) bolesnika koji ga nisu primali (328±14 (t1) naspram 322±18 mol/L). Bolesnici koji su primali karvedilol i oni koji nisu imali su isti nivo CPK na početku hirurške procedure (t1) (78±6 naspram 83±13 U/L) ali je na kraju procedure uočen niži porast aktivnosti CPK (p<0.05) u bolesnika koji su primali karvedilol u poređenju sa bolesnicima koji nisu (338±46 prema 644±103 U/L) (t2). Ovakvi rezultati sugerišu da intervencija na otvorenom srcu dovodi do povećanja nivoa CPK, ali je taj efekat bio manje izražen u bolesnika koji su imali viši nivo UA. Zaključak: Naši rezultati ukazuju na moguću pozitivnu ulogu mokraćne kiseline u zaštiti od oštećenja koja može prouzrokovati reperfuzija. Povećanje nivoa UA pre operacije može predstavljati koristan faktor tokom CABG procedure u bolesnika koji su primali karvedilol, jer snižava nivo peroksinitrita kao jedan od uzroka oštećenja od reperfuzije. Naši rezultati pokazuju uticaj UA na nivoe CPK pri kraju hirurške intervencije, što pak ukazuje na to da uvećani nivoi UA mogu štititi srce od biohemijskih oštećenja izazvana peroksinitritom tokom CABG intervencije

Bioavailability and catalytic properties of copper and iron for Fenton chemistry in human cerebrospinal fluid

A breakdown in homeostasis of redox-active metals represents an important factor for neurodegeneration. We have used EPR spectroscopy and BMPO spin-trap to investigate the catalytic properties and ligand modulation of redox activity of copper and iron in human cerebrospinal fluid (CSF). In contrast to iron, copper supplementation provoked a statistically significant increase in hydroxyl free radical generation in CSF treated with H(2)O(2). However, in a binary copper/iron containing Fenton system, iron catalytically activated copper. The chelator EDTA, which represents a model of physiological metal ligands, completely prevented copper's redox activity in CSF, while iron chelation led to a significant increase in hydroxyl radical generation, indicating that copper and iron do not only have diverse catalytic properties in the CSF but also that their redox activities are differently modulated by ligands. The application of DDC reduced hydroxyl radical generation in the CSF containing catalytically active metals (free Cu(2+) or Fe(3+)-EDTA complex). We conclude that chelators, such as DDC, are capable of preventing the pro-oxidative activity of both metals and may be suitable for reducing hydroxyl radical formation in certain pathophysiological settings

Glutathione status in the blood of rats after reticulocytosis induced by phenylhydrazine and bleeding

In this experiment, we compared the in vivo effects of phenylhydrazine (PHZ) and bleeding treatment on the redox status and glutathione antioxidative mechanism parameters in the plasma and red blood cells (RBC) of rats. Results showed a lower level of reactive oxygen species (ROS), a higher level of lipid peroxidation and the effective antioxidative role of the glutathione system in the blood of bleeding rats. PHZ-treatment induced higher concentrations of ROS and an accumulation of oxidized glutathione in the plasma, while the glutathione system showed a satisfactory antioxidative capacity in the RBC of rats. When comparing the two anemic groups, the PHZ-treated rats showed marked oxidative stress in the plasma.Cilj ovog rada je bio da se isprate in vivo efekti fenilhidrazinom (PHZ) i krvarenjem indukovane retikulocitoze na parametre redoks i glutationskog antioksidativnog statusa u plazmi i crvenim krvnim ćelijama (RBC) pacova. Rezultati pokazuju niži nivo reaktivnih vrsta kiseonika (ROS), viši nivo lipidne peroksidacije i efikasnu antioksidacionu ulogu glutationskog sistema u krvi iskvavljenih pacova. Tretman PHZom prouzrokovao je više koncentracije ROSa i akumulaciju oksidovanog glutationa u plazmi, dok je glutationski sistem pokazao efikasan antioksidativni kapacitet u RBCu pacova. Kada se uporede dve anemične grupe, izraženiji oksidacioni stres se javlja u plazmi pacova tretiranih PHZom.Projekat ministarstva br. 143035

Redox imbalance in peripheral blood of type 1 myotonic dystrophy patients

Objectives: The aim of our study was to determine if redox imbalance caused by the activities of antioxidant enzymes existed in erythrocytes of type 1 myotonic dystrophy ( DM1) patients. Methods: The activities of erythrocyte superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase were measured in 30 DM1 patients and 15 healthy controls ( HCs). The obtained values were correlated with the Muscular Impairment Rating Scale ( MIRS) score and creatine kinase ( CK). Results: Superoxide dismutase and catalase activities were lower in DM1 patients compared to HCs. A positive correlation was found between disease duration and MIRS score as well as with glutathione reductase activity. In DM1 patients, there were positive correlations between catalase, glutathione peroxidase, and glutathione reductase activities. After sub-dividing DM1 patients according to CK levels, superoxide dismutase activity was still statistically different from HCs. However, catalase activity was significantly lower only in DM1 patients with increased CK. Discussion: Undesirable alterations in antioxidant enzyme activities during DM1 disease progression may result in conditions favoring oxidative stress and changes in metabolism which together could contribute to muscle wasting

Original scientific paper

Introduction of the interdependence between the glutathione half-cell reduction potential and thermodynamic parameter