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A V Kozlov

Publications and source records attributed to A V Kozlov.

At least 37 records · Page 2Linked to original sources
PubMed1999

Nitrite reductase activity is a novel function of mammalian mitochondria.

Nitrite, which is the major stable degradation product of nitric oxide, exists in all tissues capable of nitric oxide synthesis from L-arginine. The present study provides experimental evidence that nitrite in contact with respiring mitochondria accepts reducing equivalents from the ubiquinone cycle of the respiratory chain. Univalent reduction of nitrite was totally inhibited by myxothiazol. We therefore conclude on the involvement of redox cycling that ubisemiquinone is associated with the bc1 complex. Recycling of nitric oxide degradation products via these electron carriers may become a threat to energy-linked respiration since nitric oxide in direct contact with mitochondria was shown to slow the energy-linked respiration down and to trigger a mitochondrial source for superoxide radicals. Until now, the existence of nitrite reductase activity was only demonstrated in plants and bacteria. In addition, the present observation elucidates the existence of a nitric oxide synthase-independent nitric oxide source.

Animals↗
PubMed1999

Dihydrolipoic acid maintains ubiquinone in the antioxidant active form by two-electron reduction of ubiquinone and one-electron reduction of ubisemiquinone.

Dihydrolipoic acid (DHLA) is a constituent of cellular energy metabolism, where it cycles between the oxidized and reduced form. The two thiol residues of DHLA make this biomolecule susceptible to most radical species and prevent Fenton-type reactions by chelating free iron. In this study we present a novel mode of action by which DHLA exerts antioxidant function in combination with coenzyme Q (ubiquinone). DHLA was found to reduce ubiquinone to ubiquinol by the transfer of a pair of electrons, thereby increasing the antioxidant capacity of coenzyme Q in biomembranes. In addition, ubisemiquinone, which was earlier shown to be an active oxygen radical source when existing in the anionic form, is removed from equilibrium by the addition of a single electron from DHLA. The high reactivity of DHLA with this potentially deleterious ubisemiquinone species not only prevents the formation of prooxidants, it also keeps ubiquinone in its antioxidant active form. Experimental data of this study demonstrate a superadditive effect of ubiquinone in combination with DHLA in preventing peroxidation of biomembranes.

Animals↗
PubMed1999

Critical aspects of the antioxidant function of coenzyme Q in biomembranes.

Coenzyme Q (ubiquinone, UQ) is increasingly considered as a significant natural antioxidant, which protects biomembranes in concert with alpha-tocopherol. In vitro experiments demonstrated that reduced UQ (ubiquinol) can improve the chain-breaking activities of alpha-tocopherol by recycling the antioxidant-derived reaction product, the chromanoxyl radical, to the native antioxidant. Less attention, however, was devoted to the antioxidant-derived reaction products of reduced UQ. Although both alpha-tocopherol and ubiquinol were found to be equally effective in scavenging chain-propagating lipid radicals. alpha-tocopherol protected lipid membranes from lipid peroxidation more efficiently than ubiquinol. The present study not only provides data which document this discrepancy but also contributes experimental data on the existence of ubiquinol derived pro-oxidants, which give an explanation of this phenomenon.

Antioxidants↗
PubMed1999

Nitric oxide synthase-independent generation of nitric oxide in rat skeletal muscle ischemia-reperfusion injury.

We have used electron paramagnetic resonance to investigate the time course of nitric oxide (NO) generation and its susceptibility to inhibitors of nitric oxide synthase (NOS) in ischemia-reperfusion (IR) injury to rat skeletal muscle in vivo. Significant levels of muscle nitroso-heme complexes were detected 24 h postreperfusion, but not after at 0.05, 3, and 8 h of reperfusion. The levels of muscle nitroso-heme complexes were not decreased by the NOS inhibitor N-nitro-L-arginine methyl ester as a single dose (30 mg/kg) prior to reperfusion or as multiple doses continued throughout the reperfusion (total administered, 120 mg/kg) or by the potent NOS inhibitor S-methylisothiourea (3 mg/kg). In contrast, nitroso-heme levels were reduced by the glucocorticoid dexamethasone (2.5 mg/kg). Muscle necrosis in vitro did not result in the formation of nitroso-heme complexes. The finding that reperfusion after ischemia is necessary for NO formation suggests that an inflammatory pathway is responsible for NOS-independent NO formation in IR injury to skeletal muscle.

Animals↗
PubMed1999

Nitric oxide and nitric oxide synthase in the early phase of perinatal asphyxia of the rat.

The role of nitric oxide, a compound involved in neurotransmission and regulation of cerebral blood flow, in cerebral ischemia is still not fully elucidated yet. Although well studied in adult systems of cerebral ischemia/hypoxia, information on nitric oxide in perinatal asphyxia is limited and, in particular, no direct evidence for its generation has been provided. We therefore decided to study nitric oxide generation in brain of asphyctic rat pups by biophysical and biochemical methods. We used a simple, non-invasive rat model resembling the clinical situation in perinatal asphyxia: rat pups delivered by Caesarean section were placed into a water bath at 37 degrees C still in patent membranes for various asphyctic periods (up to 20 min). Brain pH, cerebral blood flow, neuronal nitrix oxide synthase messenger RNA (by northern and dot blot analysis), immunoreactive protein (by western blot analysis) and nitric oxide synthase activity were determined; generation of nitric oxide was evaluated directly by electron paramagnetic resonance spectroscopy. Neuronal nitric oxide synthase messenger RNA activity and nitric oxide generation were unaffected, whereas neuronal nitric oxide synthase-immunoreactive protein of 150,000 mol. wt was decreased and of 136,000 mol. wt was increased with the length of the asphyctic period. This is the first report on direct evidence for the generation of nitric oxide in perinatal asphyxia and we demonstrate that nitric oxide production remains unaffected even by 20 min of asphyxia, at a time-point when cerebral blood flow was increased four-fold and severe acidosis was present. However, it was found that levels of immunoreactive neuronal nitric oxide synthase of 136,000 mol. wt were increased paralleling the length of asphyxia. Levels of the 150,000 mol. wt immunoreactive neuronal nitric oxide synthase protein decreased, suggesting a different regulation pattern. Thus, the present biochemical and biophysical results form the basis for further investigations on nitric oxide in perinatal asphyxia.

Animals↗
PubMed1998

Antioxidant-derived prooxidant formation from ubiquinol.

Ubiquinol (QH2) is increasingly used as antioxidant for the treatment of a variety of diseases and the modulation of biological aging; however, the biological significance of secondary reaction products has been disregarded so far. Our studies on the antioxidant activity of ubiquinol in peroxidizing lipid membranes demonstrate the existence of ubisemiquinone (SQ*) as the first reaction product of ubiquinol. A fraction of SQ* derived from the antioxidative activity of QH2 was detected in the outer section of the membrane bordering the aqueous phase. This localization allows an access of protons and water from the aqueous phase to SQ* a prerequisite earlier found to trigger autoxidation. Superoxide radicals emerging from this fraction of autoxidizing SQ* form H2O2 by spontaneous dismutation. SQ* not involved in autoxidation may react with H2O2. Transfer of the odd electron to H2O2 resulted in HO* and HO- formation by homolytic cleavage. An analogous reaction was also possible with lipid hydroperoxides which accumulate in biological membranes during lipid peroxidation. The reaction products emerging from this reaction were alkoxyl radicals. Both HO* and alkoxyl radicals are strong initiators and promoters of lipid peroxidation. Indirect evidence of the existence and prooxidative activities of these secondary reaction products came from comparative studies with vitamin E. While in the absence of other reactants, QH2 and vitamin E were equally effective in scavenging lipid radicals; the radical protecting activity of QH2 was found to be significantly lower as compared to vitamin E when these antioxidants operate in peroxidizing lipid membranes. This discrepancy reveals that the antioxidative activity of coenzyme Q is compulsorily linked to the formation of split products counteracting the membrane protective effect of this natural antioxidant.

Antioxidants↗
PubMed1997

Air blast-induced pulmonary oxidative stress: interplay among hemoglobin, antioxidants, and lipid peroxidation.

Blast overpressure (BOP) is a phenomenon that describes the instantaneous rise in atmospheric pressure above ambient, resulting from the firing of large caliber weapons or from military or civilian explosions. Exposure to BOP results in injury to the gas-filled organs, such as the lungs, which exhibit a contusion-type injury. We examined the effects of BOP in rats at 5 and 60 min after exposure to a low-level BOP (62 +/- 3 kPa). The exposure was found to cause oxidative stress in the lung that was characterized by 1) a 3.5-fold decrease in total antioxidant reserves, 2) a depletion of the major water-soluble antioxidants ascorbate and glutathione (GSH) by 50 and 75%, respectively, 3) a depletion of lipid-soluble antioxidant vitamin E by 30%, 4) a 2.5-fold increase of fluorescent end products of lipid peroxidation, and 5) an increased methemoglobin (metHb) content at 60 min after exposure. To elucidate the role of released hemoglobin (Hb) in blast-induced oxidative stress, we studied the interactions of oxyhemoglobin (oxyHb), metHb, and the oxoferryl from of Hb free radical species with two physiologically important reductants, ascorbate and GSH. We found that both ascorbate and GSH were able to convert oxyHb to metHb in a reaction that yielded the one-electron oxidation intermediates semidehydroascorbyl radical and glutathionyl radical, respectively. This reaction did not occur under anaerobic conditions, suggesting that oxyHb-bound O2 acted as the electron acceptor. OxyHb induced peroxidation of cis-parinaric acid in the presence but not absence of ascorbate or GSH. Thus the prooxidant action of water-soluble antioxidants via redox cycling of oxyHb and metHb may promote oxidative stress rather than prevent it.

Animals↗
PubMed1996

'Free' iron, as detected by electron paramagnetic resonance spectroscopy, increases unequally in different tissues during dietary iron overload in the rat.

'Free' iron concentration, as determined by electron paramagnetic resonance (EPR) spectroscopy, and lipid peroxidation (LPO), as determined by thiobarbituric acid test, were assessed in the lung, heart, liver, spleen, brain and kidney of rats subjected to experimental iron overload. Two tests, Desferal- and NO-available iron, were used to measure 'free' iron and gave comparable results. The most pronounced accumulation of 'free' iron was observed in liver, kidney and spleen. Differences between control and iron loaded animals increased during the initial 90 days of treatment. Between 90 and 180 days 'free' iron concentration reached a steady state level, or even decreased, as in the case of liver. Lipid peroxidation level, measured in the organs of both treated and matched controls, did not give any significant difference during the initial 90 days of treatment. A significant augmentation was observed in liver, kidney, spleen and heart at 180 days. The results of the present research show that, under conditions of moderate siderosis, the occurrence of LPO is partially related to the level of 'free' iron.

Animals↗
PubMed1996

Decreased transferrin and increased transferrin saturation in sera of women with preeclampsia: implications for oxidative stress.

OBJECTIVE: The concerted iron-binding antioxidant activity of transferrin and ceruloplasmin decreases with increasing transferrin saturation by iron. We examined interactions between serum iron and iron-binding capacity and concentrations of the lipid peroxidation metabolite malondialdehyde in normal and preeclamptic pregnancies. We also asked if the release of iron from free hemoglobin by lipid hydroperoxides is a potential mechanism to increase transferrin saturation in preeclampsia. STUDY DESIGN: Predelivery and 24 to 48 hour postpartum venous blood was collected from 19 women with uncomplicated pregnancies and 17 with preeclampsia. Serum iron, iron binding capacity, and malondialdehyde were measured. In a subset of predelivery samples electron paramagnetic resonance spectroscopy was used to determine diferric transferrin, total transferrin, and ceruloplasmin concentrations and to examine interactions of an organic hydroperoxide with hemoglobin and transferrin. RESULTS: Antepartum serum iron concentrations were 46% greater, percent saturation of iron binding capacity was 98% greater, and malondialdehyde 50% greater, whereas total iron-binding capacity was 14% lower, in women with preeclampsia. By 48 hours post partum group differences between these variables other than total iron-binding capacity were not observed. Electron paramagnetic resonance spectroscopy confirmed antepartum differences and that total iron-binding capacity and percent saturation were equivalent to total transferrin and the ratio diferric transferrin/total transferrin, respectively. Antepartum concentrations of ceruloplasmin were not different. Antepartum malondialdehyde concentrations correlated positively with percent transferrin saturation and negatively with unsaturated iron-binding capacity (apotransferrin). Electron paramagnetic resonance spectroscopy demonstrated that the release of iron from free hemoglobin by lipid hydroperoxides in serum is a potential mechanism to increase transferrin saturation. CONCLUSION: Increased transferrin saturation and decreased unsaturated iron-binding capacity in preeclampsia may occur consequent to oxidative stress and then further promote oxidative stress by decreasing serum antioxidant buffering against redox-active iron.

Adult↗
PubMed1995

Ex vivo demonstration of nitric oxide in the rat brain: effects of intrastriatal endothelin-1 injection.

Nitric oxide (NO) is a novel transmitter with multiple functions in endothelium and neuronal tissue. In particular, it has been implicated in the pathogenesis of neurodegenerative diseases. The aim of the present study was to demonstrate the ex vivo detection of NO in basal conditions and after ET-1 intrastriatal injection by means of electron paramagnetic resonance (EPR) spectroscopy using locally injected hemoglobin (Hb) as a NO trapping agent. The extent of neostriatal damage after Hb and ET-1 injections was assessed by means of immunocytochemistry with a monoclonal antibody against dopamine and cAMP-phosphoprotein M(r) 32 (DARPP-32), which is considered a marker of striatal intrinsic neurons. In the absence of local Hb injection, no signal related to endogenous NO was detected in the neostriatum, suggesting that endogenous NO trapping agents are not sufficiently concentrated to allow NO detection with the present technique. Instead, 1 h after Hb injection, a clear nitrosyl-Hb signal can be detected in neostriatal homogenates. ET-1, a powerful vasoconstrictor agent, was used to cause neuronal loss in the neostriatum. No change in nitrosyl-Hb signal was observed in neostriatal 1 h after ET-1 injection, whereas an almost 3-fold increase in the signal intensity was present 24 h after ET-1 injection. The analysis of neostriatal damage showed that Hb injection did not cause either significant damage of striatal tissue or potentiation of ET-1-induced lesions. In conclusion, the present technique allows ex vivo detection of NO in the brain. The delayed increase in NO observed after ET-1 injection indicates that this molecule may participate in the development of slowly progressive neuronal damage occurring at late post-ischemic times.

Animals↗
PubMed1995

[Analysis of a chromosomal DNA fragment isolated from a hybrid autonomous plasmid of Chlamydomonas reinhardtii].

The ability of a previously cloned fragment of nuclear DNA of Chlamydomonas reinhardtii (a 2.4-kb BamH 1-EcoR I fragment) to function as the origin of replication of plasmid pARG7.8.2.4, carrying the homologous arg7 marker gene of prototrophy or arginine in cells of C. reinhardtii transformants, was studied. As shown by Southern blotting, plasmid pARG7.8.2.4 integrates at several sites of chromosomal DNA of chlamydomonas without autonomous maintenance in cells. Results obtained using Southern blotting and plasmid rescue techniques demonstrated that plasmid pARG7.8, containing only the arg7 gene, is able both to integrate into the chromosome and to be maintained, in some cases, in the autonomous state in Arg+ transformants of alga. Analysis of the nucleotide sequence of the DNA fragment under study revealed several types of GC-enriched repeats that manifested homology with the nucleotide sequence of the arg7 gene in introns 10 and 12.

Animals↗
PubMed1993

Simultaneous determination of Fe(III) and Fe(II) in water solutions and tissue homogenates using desferal and 1,10-phenanthroline.

At neutral pH values 1,10-phenanthroline forms a colored complex with Fe(II), but it does not form such a complex with Fe(III). On the contrary, only Fe(III) forms with desferal a yellow complex with a g = 4.3 electron paramagnetic resonance (EPR) signal, but Fe(II) is rapidly oxidized by desferal to Fe(III), which gives then a yellow complex. On the basis of these facts, a method for simultaneous determination of both Fe(II) and Fe(III) ions was elaborated using a desferal-phenanthroline mixture. Two ways of detecting Fe(II) and Fe(III) have been suggested: (1) the spectrophotometric method for transparent water solutions, and (2) the EPR-spectrometric method for turbid solutions and tissue homogenates. The latter method was applied for determination of free and weakly bound iron in rat liver. The Fe(II) amount in intact liver was 22.2 +/- 7.6 nmol/g of wet tissue; free Fe(III) was not found.

Animals↗
PubMed1993

[Ways to improve ambulatory care for patients with surgical infection].

A deep analysis of purulent surgical diseases and purulent complications in 5,780 out-patients was conducted by means of a specially worked out punch card. Four groups were identified posttraumatic purulent complications--accidental wounds, abscesses, phlegmons, panaris, regional lymphadenitis, osteomyelitis, etc.--3,274 (56.6%); purulent diseases--furuncle, carbuncle, suppurating atheroma, hidradenitis, etc.--2,208 (38.2%); postoperative purulent complications--155 (2.7%), and postinjection purulent complications--143 (2.5%). The prevailing therapeutic measures were surgery and antiseptic agents used during treatment (for irrigation and dressings) of the wounds; the efficacy of these agents was of great significance. The effect of three antiseptics was studied: nitrofurazone 1:5000, dimexid, and electrochemically activated water with pH-2.4. The two first antiseptics were applied both in the first and second phases of inflammation, activated water with pH 2.4 was used only in the first phase, and activated water with pH 11.3 was used in wound dressing in the second stage of healing. The average duration of temporary incapacity was 19.7 +/- 3.2 days in 2,661 patients treated with 1:5000 nitroturazone, 16.2 +/- 2.2 days in 2,700 patients treated with dimexid, and 10.3 +/- 1.4 days in 419 patients treated with activated water. Depending on the methods of wound treatment, the economical losses per patient were 1,380, 982, and 624 roubles, respectively.

Adult↗
PubMed1992

Immunoelectronmicroscopic study of the nucleoid structure of hydrogen bacteria.

Electron microscopical studies of the nucleoid structure of hydrogen bacteria using ultrahin sections and spread DNA from bacterial cell lysates revealed a different DNA packaging in the cell. A compact state of the major part of DNA at all growth stages and stability of nucleosome-like structures were shown. The use of antibodies to HU protein of E. coli labelled by protein A-colloidal gold demonstrated the immunological relationship between HU protein of E. coli and histone-like proteins of Alcaligenes eutrophus and their possible role in the nucleosome-like DNA packaging in procariotic genome.

Alcaligenes↗
PubMed1992

Intracellular free iron in liver tissue and liver homogenate: studies with electron paramagnetic resonance on the formation of paramagnetic complexes with desferal and nitric oxide.

Treatment of intact liver and liver homogenate with sodium nitrite, or desferal, brings about the appearance of g = 2.03 and g = 4.3 electron paramagnetic resonance spectroscopy (EPR) signals, respectively. The g = 2.03 signal is conditioned by the formation of dinitrosyl complexes of Fe(II); the g = 4.3 signal is related to the appearance of paramagnetic desferal-Fe(III) complexes. Desferal and sodium nitrite were administered successively into liver homogenate, resulting in only a g = 4.3 EPR signal. And, vice versa, if desferal was administered after sodium nitrite, there appeared only the signal with g = 2.03. These data testify to the fact that one and the same endogenous free iron is included in both paramagnetic centers. The concentration of iron ions was measured in intact tissue according to the formation of dinitrosyl-iron complexes and desferal-iron complexes. It was 33.2 +/- 4.6 and 20.3 +/- 4.0 nmol/g of tissue weight, respectively. The data obtained testify to the fact that free endogenous iron is present in intact tissue. Possibilities of the EPR method for estimation of the content of intracellular free iron are discussed.

Animals↗