Experimental treatment of recurrent otitis externa.

The aim of this research is to determine the effect of the hybrid material based on polyvinyl alcohol and silver nanoparticles (PVA/AgNps) in the treatment of the otitis externa as an additional component in the commercial product "Betazon Trio". It was established that the experimental creamy formula with silver concentration 600 mg/L is suitable for recovery of the microbial homeostasis when it is administrated once daily in dose 1 ml over a period of 14 days.

Gastric mucosal lesions in influenza virus infected mice: role of gastric lipid peroxidation.

The present study provides direct experimental proof that the combination of influenza virus infection A/Aichi/2/68 (H3N3) with different models of oxidative stress, such as immobilization, cold and cold-restraint, is associated with graduated oxidative disturbances in the stomach of mice, despite the absence of virus replication and inflammation in this tissue. It was found that experimental influenza virus infection is accompanied with significant changes in gastric mucosal integrity, as well as an increase in the products of lipid peroxidation in the stomachs of mice. Preliminary exposure of mice to immobilization stress and subsequent inoculation of influenza virus did not significantly influence gastric ulceration or lipid peroxidation compared with infected mice. Cold stress resulted in a significant decrease in the index of stomach ulceration and did not influence the fluorescent products of lipid peroxidation and MDA compared with infected animals. The simultaneous application of cold-restraint stress and influenza virus infection provoked synergism in the activity of all factors on the parameters under investigation. Ulceration increased approximately two-fold, as did the amount of fluorescent products of lipid peroxidation and MDA, compared with influenza virus-infected and non-stressed animals.

Effect of vitamin E and vitamin C combination on experimental influenza virus infection.

Successful antioxidant treatment of the so-called "free radical diseases" has been reported in the literature. In this study we examined the preventive effect of vitamin E and vitamin C, alone and in combination, on the damage caused by influenza virus infection (IVI). Male mice (ICR), infected with influenza virus A/2/68/(H3N2) (1.5 of LD(50)), were administered single once-daily doses of vitamin E (60 mg/kg b.w.) and vitamin C (80 mg/kg b.w.) intraperitoneally (3 days before virus inoculation). On the 5th and 7th day, respectively, after virus inoculation, animals were decapitated. Monooxygenase enzyme activity (ethylmorphine N-demethylase, amidopyrin N-demethylase, analgin N-demethylase, aniline hydroxylase, cytochrome P-450 content and NADPH-cytochrome C reductase [CCR]) was determined in liver 9000 x g supernatant. Primary and secondary products of lipid peroxidation (LPO; conjugated dienes [CD] and TBA-reactive substances) were measured in blood plasma, lung and liver 9000 x g supernatant. Vitamin E effectively restored LPO-levels increased by IVI. The effect of vitamin C was similar, but slighter. The combination (vitamin E + C) had greater effect on LPO levels than their separate administration. P-450-dependent monooxygenase activity was significantly restored and more pronounced cytochrome P-450 content and NADPH-CCR activity was noted. The preventive effect of vitamin E was stronger than the effect of vitamin C, but the combination (vitamin E + C) had the strongest effect. The superior protective effect of the combination is probably due to vitamin C's repairing effect on vitamin E's tocopheroxyl radical.

Effect of immobilization, cold and cold-restraint stress on liver monooxygenase activity and lipid peroxidation of influenza virus-infected mice.

The present study provides a direct experimental evidence that the combination of influenza A/Aichi/2/68 (H3N2) infection with different models of "oxidative stress", such as immobilization, cold and cold-restraint, is associated with graduated oxidative disturbances in the liver of mice, despite the absence of virus and inflammation in this tissue. It was found that experimental influenza virus infection is accompanied with a significant increase of lipid peroxidation products, a decrease of natural antioxidants (vitamin E, glutathione) and cytochrome P-450, an inhibition of cytochrome c reductase and liver monooxygenases (analgin- N-demethylase and amidopyrine- N-demethylase). Immobilization and cold stress, applied separately or in combination (cold-restraint), did not influence significantly any of the analysed parameters compared to those of the control group of non-infected mice. Preliminary exposure of mice to immobilization or cold stress and subsequent inoculation of influenza virus resulted in a significant increase of lipid peroxidation products and a significant decrease of vitamin E and reduced glutathione, compared with levels in control (non-infected) animals. Compared to influenza virus-infected and non-stressed animals, the changes in all these parameters were negligible. Immobilization or cold stress, applied in combination with influenza virus infection, partially prevented the suppressive effect of influenza virus on cytochrome P-450 and liver monooxygenases. A tendency towards normalization of these parameters to the control levels was observed. However, after application of cold-restraint plus influenza virus infection, the level of cytochrome P-450 and activity of cytochrome c reductase stayed markedly lower than in infected and non-stressed animals. The activities of liver monooxygenases were slightly increased compared with those of infected and non-stressed animals, but stayed relatively low compared to control (non-infected) mice. Combination of cold-restraint and influenza virus infection resulted in a greater synergistic increase of lipid peroxidation products and a greater synergistic decrease of vitamin E and reduced glutathione compared to controls, as well as to influenza virus-infected and non-stressed animals.

Effect of vitamin E supplementation on lipid peroxidation in blood and lung of influenza virus infected mice.

The influenza virus infection (A/Aichi/2/68) was associated with development of oxidative stress in lung and blood of mice, accompanied by an increase in levels of lipid peroxidation products (conjugated dienes and total malondialdehyde) and a decrease in endogenous amounts of natural antioxidant vitamin E. These effects were most pronounced on the 5th day after virus inoculation, in comparison with those on the 7th. Supplementation of mice with exogenous vitamin E before virus inoculation lead to lung and blood protection against lipid peroxidation. A marked decrease in lipid peroxidation products and an increase in vitamin E content was established in blood and lung on the 5th and 7th day after virus inoculation. The stabilizing effect of vitamin E is dose-dependent in blood and dose-independent in lung, and was most pronounced on the 5th day after virus inoculation in comparison with the 7th day.

[Studies on restraint ulcers in rats in conditions of acute copper deficiency]. / Studovi imobilizatsionni iazvi pri plukhove v usloviiata na ostur meden defitsit.

In the present experiment, we have investigated the impact of the acute copper deficiency on the pathogenesis of stress ulcers' formation in rats during cold-restraint stress. A powdered milk diet, lasting for 5 days, causes a significantly decrease of stomach copper content in rat, comparing to content in rats receiving standard laboratory diet. The severity of mucosal disturbances, expressed as total number and area of stress ulcers, is most prominent in animals receiving hypocupric diet. Their treatment with Ranitidine extends the morphological disturbances in the stomach mucosa and produces highest level of lipid peroxidation and lowest activity of superoxide dismutase in the stomach compared to the other groups. In conclusion, the appropriate copper balance plays an essential role for the natural resistance of the stomach mucosa and Ranitidine, in conditions of acute copper deficiency, makes deeper the disturbances produced by stress.

Influence of acute copper deficiency, cold-restraint stress and the H2 blocker ranitidine on the severity of acute gastric mucosal lesions and lipid peroxidation in rats.

Acute copper deficiency produces disturbances in the microcirculation and structure of extracellular matrix proteins, causes an increase in mast cell population, which is followed by an increased content of their degranulation products, produces disturbances in histamine metabolism and decreases the activity of some antioxidant enzymes. These pathogenic mechanisms are similar to the processes underlying stress ulcer formation. The histamine H2-receptor antagonist ranitidine, a drug with the highest application for stress ulcer prophylaxis, has the ability to helate the copper ion and to influence its tissue distribution and the processes of generation and neutralization of reactive oxygen species (ROS). In order to determine the interrelation between the disturbances of copper homeostasis, stress ulcers and ranitidine, we investigated the impact of a short-term diet with powdered milk in combination with cold-restraint stress with or without ranitidine on the severity of acute gastric mucosal lesions, copper content, lipid peroxidation and the activity of superoxide dismutase and catalase in the stomachs of rats.

Antioxidant properties of rimantadine in influenza virus infected mice and in some model systems.

Influenza virus infection is associated with development of oxidative stress in lung and blood plasma, viz. increase of primary and secondary lipid peroxidation products. It was established that rimantadine treatment led to a decrease of the products of lipid peroxidation in tissues of mice experimentally infected with influenza virus A/Aichi/2/68 (H3N2). The effect is strongest in blood plasma (a decrease of about 50%) and weaker in the lung (about 20%). To elucidate the mechanism of this action of rimantadine, experiments were carried out with some model systems. The capability of rimantadine to scavenge superoxide radicals (scavenging properties) was studied in a system of xanthine-xanthine oxidase to generate superoxide. The amount of superoxide was measured spectrophotometrically by the NBT-test and chemiluminesce. Rimantadine does not show scavenging properties and its antioxidant effect observed in vivo, is not a result of its direct action on the processes of lipid peroxidation and/or interaction with antioxidant enzymes. The antioxidant properties of rimantadine were investigated by measurement of induced lipid peroxidation in a Fe2+ and (Fe2+ - EDTA) system with an egg liposomal suspension. Our findings with model systems do not prove an antioxidant or prooxidant effect of the drug on the processes of lipid peroxidation. Apparently, the observed antioxidant effect of rimantadine in vivo is not connected directly with free radical processes in the organism.

Determination of malondialdehyde in biological samples by solid-phase extraction and high-performance liquid chromatography.

An analytical procedure for determination of malondialdehyde in tissue homogenates and blood serum was developed. A reaction with 2,4-dinitrophenylhydrazine is used followed by cleaning up of the derivative by solid-phase extraction. The samples were analyzed by isocratic high-performance liquid chromatography (HPLC) using a narrow-bore HPLC-column. A good separation of the 1-pyrazole peak from that of 2,4-dinitrophenylhydrazine was observed. A high linear dependence was established by the concentration of 1-pyrazole in the range of 10-5000 ng/ml. The detection limit of the method applied for tissue homogenates and blood serum was approximately 10 ng/ml or lower, and RSD of the method was 9% (n = 8). The peak of 1-pyrazole for these samples was well separated from the other matrix peaks. Experiments carried out evaluated that the solid-phase extraction might be an effective step of the sample preparation, significantly increasing the selectivity of the analysis and the life-time of the column. The method seems to be applicable for determination of malondialdehyde in different biological samples.

Effect of vitamin E on lipid peroxidation and liver monooxigenase activity in experimental influenza virus infection.

Influenza virus infection was associated with development of oxidative stress in liver of mice, viz. increase in amount of lipid peroxidation products, decrease in cytochrome P-450 and NADP. H-cytochrome c-reductase activity, and inhibition of liver monooxygenases (aniline hydroxylase, ethylmorphine-N-demethylase, amidopyrine-N-demethylase and analgin-N-demethylase). These effects were most pronounced on the 7th day after virus inoculation as compared to the 5th one. Supplementation of mice with vitamin E before virus inoculation leads to liver protection against oxidative stress and toxicosis. A marked decrease of lipid peroxidation products and an increase of cytochrome P-450 and activities of monooxygenases was established. The stabilizing effect of vitamin E was dose-dependent and was most pronounced on the 5th day after virus inoculation as compared to the 7th one.

Pharmacodynamic of the antioxidant action of alpha-tocopherol and its derivatives in liver, brain, heart and skeletal muscles.

The aim of the present work was to determine the pharmacodynamics of antioxidant effect of alpha-tocopherol and its derivatives (alpha-tocopheryl esters and chromanols with different chain-length) in the animal tissues, as well as the role of cytochrome P-450 in biotransformation of these compounds. Alpha-tocopherol and its derivatives were injected intraperitoneally in rats or mice in a single dose of 100 mmol per kg b.w. The animals were sacrificed at different time intervals (0, 1, 2, 4, 8, 12, 24, 36 hours) and the liver, heart, brain and skeletal muscles were removed, homogenized and incubated with lipid peroxidation (LPO) inducers (Fe2+ + ascorbate). LPO was evidenced by the generated malone dialdehyde (MDA). Data were expressed as percentage of LPO inhibition by alpha-tocopherol or its derivatives as compared to control group. The kinetic curves of the inhibitory action of alpha-tocopherol and its derivatives on LPO were characterized by three phases: a phase of increasing antioxidant activity, a phase of maximal antioxidant activity (about 60-95% LPO inhibition), and a phase of decreasing antioxidant activity. Alpha-tocopheryl esters possessed dynamics of antioxidant action the same as alpha-tocopherol. Therefore the hydrolysis of alpha-tocopheryl esters in animal organism is not a limiting factor for their antioxidant effect. The alpha-tocopherol derivatives with short chain-length (C1, C6) had a shorter half-life in animal tissues as compared to alpha-tocopherol or its esters. In vitro experiments showed that C1 and C6 are substrates of cytochrome P-450. In contrast, alpha-tocopherol and its esters did not bind to cytochrome P-450 even at concentrations as high as 10 mmol/l. Apparently, C1 and C6 underwent biotransformation and were excreeted more quickly from the organism.

Effect of He-Ne laser treatment on the level of lipid peroxidation products in experimental cataract of rabbit eyes.

The effect of low-intensity laser irradiation on the processes of lipid peroxidation in lens homogenate and aqueous humor during experimental diquat-induced cataract of rabbit eyes was studied. The levels of primary and secondary products of lipid peroxidation (LPO), conjugated dienes and thiobarbituric acid reactive substances (TBARS), were evaluated. We found that the experimental cataract model leads to a significant increase in the content of conjugated dienes and in the content of TBARS both in lens homogenate and in aqueous humor. The data obtained support the important role of oxidative stress in the development of the diquat-induced cataract model. Low-intensity laser treatment does not provoke a significant decrease in conjugated dienes or in TBARS in either lens homogenate or aqueous humor. Although our therapeutic scheme led to a slightly decreased level of LPO products, we conclude that the effect of low-intensity laser-irradiation may depend on the dose applied, individual tissues and other factors.

Specificity of glycosaminoglycan suppression of endothelin-1 production by human umbilical vein endothelial cells.

Endothelin-1 (ET-1) is the most potent vasoconstrictor peptide found in nature. Its production is stimulated by thrombin. By inhibiting thrombin we have previously shown that heparin, a highly negatively-charged glycosaminoglycan (GAG), suppresses the production of ET-1 by cultured human umbilical vein endothelial cells (HUVEC). The purpose of our study is to determine the effect of other GAGs and related compounds on ET-1 production. The GAGs and related compounds used in the study were: chondroitin sulfate A, chondroitin sulfate B, chondroitin sulfate C, fucoidin, low molecular weight dextran sulfate, high molecular weight dextran sulfate, and hyaluronan. HUVEC were incubated for 48 hr with media containing these GAGs and related compounds and with media without GAG as control. ET-1 levels were measured by radioimmunoassay. GAGs and related molecules with higher sulfate content, heparin, chondroitin sulfate B, low and high molecular weight dextran sulfates significantly suppressed ET-1 production by HUVEC. Fucoidin also suppressed ET-1 production despite its lower sulfate content, probably because of its structural similarity to heparin. These compounds may be useful for future in vivo studies.

Nitric oxide products degrade chondroitin sulfates.

Nitric oxide (NO) is a potent endogenous vasodilator that is elevated in response to inflammation. Inflammation also produces high levels of superoxide, which combines with NO to produce peroxynitrite (PN). We have previously reported that NO degrades heparin and heparan sulfate under acidic conditions and that PN degrades hyaluronan (HA) at neutral pH. Heparin and HA are glycosaminoglycans (GAGs) widely distributed in the extracellular matrix of tissues. Disruption of intestinal GAGs, particularly the chondroitin sulfates, were linked to inflammatory bowel diseases. Chondroitin sulfate A (CSA), chondroitin sulfate B (CSB), and chondroitin sulfate C (CSC) are constituents of the basement membranes of many tissues, including the intestine. The purpose of this study is to determine whether the NO donor S-nitroso-N-acetylpenicillamine (SNAP) and PN can degrade chondroitin sulfates in vitro. The NO donor SNAP (2 mM, pH 4.0) or PN (5 mM, pH 7.4) was incubated for at least 1 week at 37 degrees C with CSA, CSB, or CSC. Breakdown of CSA, CSB, and CSC was assessed by gel filtration chromatography and compared with untreated controls. Percentage degradation was calculated based on the change in peak height compared to the control. SNAP treatment partially degraded CSB and CSC, whereas PN partially degraded all three chondroitin sulfates. Nitric oxide mediated degradation of GAGs, and particularly chondroitin sulfates, may be an important pathway of inflammatory tissue damage.

The nitric oxide donor S-nitroso-N-acetyl-D,L-penicillamine degrades heparan sulfate and heparin.

Nitric oxide (NO) is a powerful vascular and neural regulator. One of the breakdown products of nitric oxide is nitrite which converts to nitrous acid, a reagent routinely used for the degradation of heparin and heparan sulfate. We have recently shown that nitric oxide gas degrades heparin and heparan sulfate through a nitrous acid mechanism (Vilar et al, 1997, Biochemical Journal, 324, 473-479). The purpose of the present study is to confirm these findings using the nitric oxide donor S-nitroso-N-acetyl-D,L-penicillamine (SNAP) under conditions that are close to those found in vivo. The results show that 2 mM SNAP releases a steady-state level of nitrite of over 200 microM. This level substantially degrades heparin and heparan sulfate at a pH of up to 5.0. This reaction may be important in breakdown of the glycosaminoglycan components of the extracellular matrix during normal and pathological conditions.

[Effect of charged particles of relativistic energy on the number of chromosome aberrations in human blood lymphocytes. Dose-response relationship and the RBE of protons, deuterons and helium ions]. / Vliianie zariazhennykh chastits reliativistskikh énergii na chastoty khromosomnykh aberratsii v limfotsitakh krovi cheloveka. Zavisimost' doza-éffekt i OBE protonov, deitonov i ionov geliia.

In experiments with human blood lymphocyte culture exposed to fast protons, deuterons and helium ions the frequency and types of chromosome aberrations have been studied. Fast charged particles of a relativistic energy are shown to have a pronounced harmful effect resulting in a sharply increased formation of exchange chromosome aberrations as compared to that produced by gamma-radiation. The RBE coefficients of the particles under study have been determined: their values vary depending on the type of radiation and the tests used.

[Mutagenic effect of the cytostatic drug thaliblastine on rat bone marrow cells when administered alone and in combination with radiation]. / Issledovanie mutagennogo éffekta tsitostatika taliblastina na kletkakh kostnogo mozga krys pri razdel'nom i sochetannom s oblucheniem vozdeistviiakh.

The cytogenetic analysis was performed in the bone marrow cells of Wistar rats treated with a therapeutic dose of thaliblastine (250 mg/kg) and exposed to gamma-rays (2 Gy). Thaliblastine alone induced chromosome aberrations and polyploid cells. The latter were the result of the stathmokinetic effect of the drug. In contrast to gamma-radiation of 2 Gy thaliblastine elicited a minor mutagenic effect. The cytogenetic effect of the combined treatment is greater than the sum of the two agents delivered separately, the maximum effect of radiation and thaliblastine being exhibited on the 8th and the 12th hour, respectively. The difference between the sum of aberrations after separate treatments and the yield of aberrations after the combined treatment is due to chromatid fragments.

[Effect of a constant magnetic field and gamma radiation on the hereditary structure of somatic cells. Effect of the combination of a constant magnetic field and ionizing radiation on human lymphocytes in vitro]. / Vliianie postoiannogo magnitnogo polia i gamma-radiatsii na nasledstvennuiu strukturu somaticheskikh kletok. Vozdeistvie sochetaniia postoiannogo magnitnogo polia i ioniziruiushchei radiatsii na limfotsity krovi cheloveka in vitro.

Samples of human whole blood were exposed in CMF (field induction, 0.3 T) for 15, 30, 45, 60, 75, 90, 120, 150, 180, 240, 300 or 360 min. 15 min following exposure, the samples were gamma-irradiated in a dose of 0.0516 C/kg (137Cs) at a dose rate of 1.95 A/kg. The following chromosome aberrations were scored: deletions dicentrics, rings, and symmetrical exchanges. Exposure of the blood in CMF for 15 to 360 min decreased radiation damage to cells as compared with unexposed irradiated samples. The extention of time from 15 to 180 min increases the effect the smallest amount of chromosome damages being scored at 150-180 min. A 2.8 - fold, 3 - fold and 3.5 - fold decrease was registered in the number of aberrant cells, deletions and dicentrics, respectively. With increasing time of exposure (240 min), the radiomodifying effect started decreasing, and with 300-360 min exposure it was the same as that observed at 15-45 min.