The effect of oxidative stress on nitrosomethylurea-induced mutagenesis in sunflower Helianthus annuus L

The effect of hyberbaric oxygenation on mutagenicity of nitrosomethylurea (NMU) was examined. It was shown that in the regimes studied, hyperbaric oxygenation enhances the NMU mutagenic effect both on the nuclear and on the plastid genetic material of sunflower, but do not increase intensity of free-radical reactions and do not induce plastid mutations and chromosome aberrations per se. At high concentrations inducing chromosome aberrations (0.03%), NMU was shown to enhance the level of free-radical processes. Possible mechanisms of the increase of NMU-induced mutagenesis by hyperbaric oxygenation are discussed.     

The Influence of Hyperbaric Oxygenation (HBO) on Proliferation and Differentiation of Human Keratinocyte Cultures In Vitro

A drop in tissue oxygen partial pressure below 30mm Hg as a result of reduced perfusion in an extensive area of acute skin damage, or where a large number of chronic skin defects occur, inhibits collagen synthesis and neoangiogenesis in the various phases of wound healing. Subsequent granulation and epithelialisation are correspondingly impaired.Hyperbaric oxygenation is now recognised as a valuable supplementary method of treatment for problematic wounds. Stimulation of fibroblast and endothelial cell proliferation through Hyperbaric oxygenation has been demonstrated in numerous studies.The aim of this study was to investigate the effect of hyperbaric oxygen treatment on the proliferation and differentiation of human keratinocyte cultures.The influence of hyperbaric oxygenation on the proliferation of human keratinocyte cultures was demonstrated using flow-through cytometry and a fluorescence activated cell sorter, which detects fluorescence intensity following incorporation of 5-bromo-2-deoxyuridine in cell DNA.The degree of cell differentiation was deduced from the expression of various components of the cytoskeleton, such as cytokeratin 10 and involukrin, the production of which was quantified through the determination of monoclonal antibodies against cytokeratin 10 and involukrin from measurements of fluorescence activity in a flow-through cytometer.Hyperbaric oxygenation of cell cultures in vitro did not produce a significantly higher rate of cell proliferation, so that no increase in vitality was observed.An interesting observation following exposure to hyperbaric oxygen was the marked increase in expression of both cytokeratin 10 and involukrin, as an indication of accelerated cell differentiation.     

Mutagen specificity in the induction of karyotic versus cytoplasmic respiratory deficient mutants in yeast by nitrous acid and alkylating nitrosamides

Summary In the haploid eukaryotic organism Saccharomyces cerevisiae the induction of cytoplasmic and genic (karyotic) RD mutants was studied, using nitrous acid, nitrosomethylurethane (NMU) and nitrosoimidazolidone (NIL).The cytoplasmic or genic origin of the induced RD mutants was determined by prescreening in complementation tests with ^– and wild type tester strains. Among the mutants of all three agents we could thus score the incidence of three RD mutant types: genic, suppressive and cytoplasmic (both primary and secondary). The final identification of the cytoplasmic type was only possible through tetrad analysis, performed in the cases of HNO₂ and NMU.A distinct difference in cytoplasmic versus genic mutagen specificity was observed between HNO₂ and NMU. HNO₂ was unable to induce cytoplasmic RD mutants but it proved to be highly efficient in the induction of genic RD mutants. In contrast, NMU induced more cytoplasmic effects was it possible to detect mutagenic specificities which, solely on the basis of karyotic action, were not detectable.     

Hyperbaric oxygen induces apoptosis via a mitochondrial mechanism

During therapeutic hyperbaric oxygenation lymphocytes are exposed to high partial pressures of oxygen. This study aimed to analyze the mechanism of apoptosis induction by hyperbaric oxygen. For intervals of 0.5–4 h Jurkat-T-cells were exposed to ambient air or oxygen atmospheres at 1–3 absolute atmospheres. Apoptosis was analyzed by phosphatidylserine externalization, caspase-3 activation and DNA-fragmentation using flow cytometry. Apoptosis was already induced after 30 min of hyperbaric oxygenation (HBO, P < 0.05). The death receptor Fas was downregulated. Inhibition of caspase-9 but not caspase-8 blocked apoptosis induction by HBO. Hyperbaric oxygen caused a loss of mitochondrial membrane potential and caspase-9 induction. The mitochondrial pro-survival protein Bcl-2 was upregulated, and antagonizing Bcl-2 function potentiated apoptosis induction by HBO. In conclusion, a single exposure to hyperbaric oxygenation induces lymphocyte apoptosis by a mitochondrial and not a Fas-related mechanism. Regulation of Fas and Bcl-2 may be regarded as protective measures of the cell in response to hyperbaric oxygen.     

Quasi-Adaptive Response to Alkylating Agents in <Emphasis Type="Italic">Escherichia coli</Emphasis>: A New Phenomenon

An original hypothesis of a quasi-adaptive response to nitrosomethylurea (NMU) in Escherichia coli cells was verified experimentally. In contrast to the true Ada response, which is induced in cells pretreated with a sublethal dose of NMU, a quasi-adaptive response was induced using NO-containing dinitrosyl iron complex with glutathione (DNIC_glu). Quasi-adaptation increased expression of the Ada regulon and cell resistance to the cytotoxic and mutagenic effects of NMU. The levels of alkA, alkB, and aidB gene expression in quasi-adaptation were higher than in the true Ada response. Thus, experimental evidence was obtained for the alternative mechanism regulating the function of the Ada sensory protein in controlling expression of the Ada regulon during the adaptive response. The free iron—chelating agent o-phenanthroline (OP) facilitated degradation of DNIC_glu (by electron paramagnetic resonance (EPR) spectra) and considerably or completely inhibited gene expression in the quasi-adaptive response. The new phenomenon extends the functional range of NO compounds to include a role in genetic signal transduction within the Ada response system in addition to similar roles in the SoxRS, SOS, and OxyR systems in E. coli.__________Translated from Genetika, Vol. 41, No. 5, 2005, pp. 607–613.Original Russian Text Copyright © 2005 by Vasilieva, Moschkovskaya.     

Effect of hyperbaric air on endotoxin from <Emphasis Type="Italic">Bacteroides fragilis</Emphasis> strains

The aim of the project was to determine any effect of hyperbaric air on Bacteroides fragilis strains cultivated under hyperbaric conditions. Previously, it was hypothesized that there was a correlation between the presence of Bacteroides bacteria in patients preferring a meaty diet and cancer of the small intestine, and particularly of the large intestine and rectum. With respect to the fact that Bacteroides fragilis (BAFR) group are important producers of endotoxins, measurement and statistical evaluation of endotoxin production by individual strains of isolated Bacteroides species were used to compare bacteria isolated from various clinical samples from patients with colon and rectum cancer in comparison with strains isolated from other non-cancer diagnoses. Endotoxin production was proven by quantitative detection using the limulus amebocyte lysate (LAL) test in EU/mL. Production of endotoxins in these bacteria cultured under hyperbaric air conditions was higher than those strains cultured under normobaric anaerobic conditions. But these differences in endotoxin production were not statistically significant (t test with log-transformed data, p value = 0.0910). Based on a two-tier t test for lognormal data, it is possible to cautiously conclude that a statistically significant difference was found between endotoxin production by Bacteroides fragilis strains isolated from non-carcinoma diagnoses (strains (1–6) and strains isolated from colorectal carcinoma diagnoses (strains 7–8; Wilcoxon non-parametric test p = 0.0132; t test = 0.1110; t test with log-transformed data, p value = 0.0294).     

Adaptogenic effects of furostanol glycosides of <Emphasis Type="Italic">Dioscorea deltoidea</Emphasis> wall on oxidative processes in tomato plants during biotic stress

The effect of furostanol glycosides of a cell culture of Dioscorea deltoidea Wall on oxidative processes in tomato plants subjected to invasion with the gall nematode Meloidogyne incognita Kofoid et White was studied. We showed that furostanol glycosides induce a nonspecific defensive response in plants. Exposure of cell membranes to furostanol glycosides causes rearrangements in fatty acids, resulting in the formation of conjugated dienes, which makes molecules thermodynamically more stable under stress conditions. The study of changes in the activity of peroxidases of intact plants and plants affected with the nematode, which were treated with furostanol glycosides, showed that the protective effect of the guaiacol-dependent peroxidase is more long-term than the effect of the benzidine-dependent peroxidase.__________Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 3, 2005, pp. 347–353.Original Russian Text Copyright © 2005 by Vasileva, Vanyushkin, Zinoveva, Udalova, Volkova, Nosov, Paseshnichenko.     

SkQ1 Controls <Emphasis Type="Italic">CASP3</Emphasis> Gene Expression and Caspase-3-Like Activity in the Brain of Rats under Oxidative Stress

Here, we studied the effect of the mitochondria-targeted antioxidant SkQ1 (plastoquinone cationic derivative) on the CASP3 gene expression and caspase-3 activity in rat cerebral cortex and brain mitochondria under normal conditions and in oxidative stress induced by hyperbaric oxygenation (HBO). Under physiological conditions, SkQ1 administration (50 nmol/kg, 5 days) did not affect the CASP3 gene expression and caspase-3-like activity in the cortical cells, as well as caspase-3-like activity in brain mitochondria, but caused a moderate decrease in the content of primary products of lipid peroxidation (LPO) and an increase in the reduced glutathione (GSH) level. HBO-induced oxidative stress (0.5 MPa, 90 min) was accompanied by significant upregulation of CASP3 mRNA and caspase-3-like activity in the cerebral cortex, activation of the mitochondrial enzyme with simultaneous decrease in the GSH content, increase in the glutathione reductase activity, and stimulation of LPO. Administration of SkQ1 before the HBO session maintained the basal levels of the CASP3 gene expression and enzyme activity in the cerebral cortex cells and led to the normalization of caspase-3-like activity and redox parameters in brain mitochondria. We hypothesize that SkQ1 protects brain cells from the HBO-induced oxidative stress due to its antioxidant activity and stimulation of antiapoptotic mechanisms.     

Natural conditions inducing programmed cell death in the yeast <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Although yeasts have been extensively used as an experimental model to study apoptosis, it is still unclear why a unicellular organism like yeast possesses a suicide program. Here we discuss three hypothetical scenarios of natural yeast suicide. We argue that by correctly deducing the physiological situation(s) for yeast to undergo cell death, one can not only improve the efficiency of yeast as model system for apoptotic studies, but also obtain a certain insight into the survival strategies of communities of organisms.Translated from Biokhimiya, Vol. 70, No. 2, 2005, pp. 323–326.Original Russian Text Copyright © 2005 by Knorre, Smirnova, Severin.This revised version was published online in April 2005 with corrections to the post codes.     

Association between defects of karyogamy and translation termination in yeast <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Mutants capable of a high frequency of cytoduction (Hfc⁺) were obtained in a haploid strain of Saccharomyces cerevisiae, suggesting impaired cytogamy. Nine of the 68 Hfc⁺ mutants showed the antisuppressor effect with respect to mutations of the SUP35 and SUP45 genes, which code for translation termination factors, or to the [PSI ⁺] factor, which is the prion form of Sup35. Cosegregation of the characters higher frequency of cytoduction and antisuppression was demonstrated for three Hfc⁺ mutants. One (HFC12-2) of the Hfc⁺ mutations exerted a dominant antisuppressor effect with respect to [PSI ⁺] and had no effect on [PSI ⁺] maintenance. On the strength of the results, an interaction was assumed for translation termination components and cytoskeleton proteins, which play a role in karyogamy in yeasts.Translated from Genetika, Vol. 41, No. 2, 2005, pp. 178–186.Original Russian Text Copyright © 2005 by Borchsenius, Repnevskaya, Kurischko, Inge-Vechtomov.     

On Antimutagenesis Mechanism in Plants

The effect of nitrosylmethylurea (NMU) on the mitotic index and the frequency of cells with aberrations, as well as the effects of pre- and posttreatment with antioxidant ambiol on the NMU effects were studied on seedlings of common winter wheat Triticum aestivum, cultivar Moskovskaya 39. Both pre- and posttreatment with ambiol resulted in antimutagenic effect but after posttreatment, the effect was lower. Irrespective of type of seedling treatment with ambiol and the time of their fixation (45, 48, and 51 h), when mitotic index is plotted versus frequency of cells with aberrations, all experimental points fall on the same regression line with coefficient of correlation of −0.82 (P < 0.001). This implies that the same mechanism underlies antimutagenic effect irrespective of when the antimutagen was applied, before or after the knockout mutagen dose. This also suggests that the antimutagenic effect is independent of the degree of the mutagen-induced damage, because by the time of posttreatment, the volume of genome damage is already determined and the antimutagen fails to change it. Finally, this suggests that irrespective of time of antimutagen treatment, the mutation frequency is reduced by the mechanism of stimulated repopulation.__________Translated from Genetika, Vol. 41, No. 5, 2005, pp. 676–679.Original Russian Text Copyright © 2005 by Serebryanyi, Zoz, Morozova.     

The role of body surfaces and ventilation in gas exchange of the abalone, <Emphasis Type="Italic">Haliotis iris</Emphasis>

The archaeogastropod Haliotis iris possesses paired bipectinate gills and normally four to six shell holes. In still water, endogenous water flow entered the branchial chamber anteriorly to the left of the head and was exhaled primarily from the three most posterior holes. The first or second anterior aperture was occasionally weakly inhalant. Cardiac interaction superimposed an oscillatory component upon ciliary ventilation but did not augment mean flow. At normal endogenous flow rates 49% of oxygen was extracted from the branchial flow, increasing to 71% at lower flows. In still water, normoxic was 0.47 μmol g⁻¹ h⁻¹. Oxyregulation occurred down to with partial oxyregulation down to 45 Torr (P _crit), and oxyconformity below this. The oxyregulatory plateau was absent in artificially ventilated animals but normoxic was higher (0.65 μmol g⁻¹ h⁻¹). Endogenous ventilation was unaffected by hypoxia to 15 Torr. Heart rate decreased by ~20% at 26 Torr before falling more steeply. Oxygen uptake from the branchial ventilation stream fully accounted for normoxic In hypoxia (<30 Torr), no uptake occurred from the head or foot despite extensive eversion of the epipodium. Blood oxygen measurements excluded the right mantle as a significant gas exchange organ. Changes in oxygen uptake caused by changes in the velocity of external water currents support the concept of induced ventilation and suggest that in still water aerobic respiration was ventilation-limited. Although ciliary ventilation appears adequate to support resting aerobic metabolism, induced ventilation may provide increased aerobic scope for activity and repayment of oxygen debt. An erratum to this article can be found at     

Polysaccharides of <Emphasis Type="Italic">Ganoderma lucidum</Emphasis>: Factors affecting their production

The conditions of polysaccharide production by the fungus Ganoderma lucidum were optimized. The maximal yield of endopolysaccharides and exopolysaccharides was observed at 25–30°C, initial pH of culture medium 4.0–6.0, and at a C:N ratio of 18:1 and 25:1, respectively. The greatest yield of mycelium was reached at a more intensive aeration, and the maximal yield of polysaccharides was observed at a less intensive aeration. The optimal ratio between fungus growth and polysaccharide production was observed at 100 rpm and an aeration of 1.0– 1.5 1/1 medium·min.Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 2, 2005, pp. 194–199.Original Russian Text Copyright © 2005 by Babitskaya, Shcherba, Puchkova, Smirnov.     

Studies of hydrolytic activity of enzyme preparations of <Emphasis Type="Italic">Penicillium</Emphasis> and <Emphasis Type="Italic">Trychoderma</Emphasis> fungi

Enzyme preparations were isolated from the culture liquid of five mutant strains of the cellulase producer Penicillium verruculosum. The hydrolytic activities of these preparations against unbleached eucalypt cellulose was compared to that of commercial preparations of Trichoderma reesei (T. longibrachiatum). In the majority of cases, P. verruculosum enzymes provided higher yields of reducing sugars (RSs) and glucose. A correlation was found between the yield of RSs and the avicelase activity of the preparations in the reaction mixture.Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 2, 2005, pp. 210–212.Original Russian Text Copyright © 2005 by Skomarovsky, Gusakov, Okunev, Soloveva, Bubnova, Kondrateva, Sinitsyn.     

Zinc ions as cytochrome <Emphasis Type="Italic">c</Emphasis> oxidase inhibitors: two sites of action

Zinc ions are shown to be an efficient inhibitor of mitochondrial cytochrome c oxidase activity, both in the solubilized and the liposome reconstituted enzyme. The effect of zinc is biphasic. First there occurs rapid interaction of zinc with the enzyme at a site exposed to the aqueous phase corresponding to the mitochondrial matrix. This interaction is fully reversed by EDTA and results in a partial inhibition of the enzyme activity (50–90%,depending on preparation) with an effective K _i of 10 µM. The rapid effect of zinc is observed with the solubilized enzyme, it vanishes upon incorporation of cytochrome oxidase in liposomes,and it re-appears when proteoliposomes are supplied with alamethicin that makes the membrane permeable to low molecular weight substances. Zinc presumably blocks the entrance of the D-protonic channel opening into the inner aqueous phase. Second, zinc interacts slowly (tens of minutes, hours) with a site of cytochrome oxidase accessible from the outer aqueous phase bringing about complete inhibition of the enzymatic activity. The slow phase is characterized by high affinity of the inhibitor for the enzyme:full inhibition can be achieved upon incubation of the solubilized oxidase for 24 h with zinc concentration as low as 2 µM. The rate of zinc inhibitory action in the slow phase is proportional to Zn²⁺ concentration. The slow interaction of zinc with the outer surface of liposome-reconstituted cytochrome oxidase is observed only with the enzyme turning over or in the presence of weak reductants, whereas incubation of zinc with the fully oxidized proteoliposomes does not induce the inhibition. It is shown that zinc ions added to cytochrome oxidase proteoliposomes from the outside inhibit specifically the slow electrogenic phase of proton transfer, coupled to a transition of cytochrome oxidase from the oxo-ferryl to the oxidized state (the F O step corresponding to transfer of the 4th electron in the catalytic cycle).Translated from Biokhimiya, Vol. 70, No. 2, 2005, pp. 160–170.Original Russian Text Copyright © 2005 by Kuznetsova, Azarkina, Vygodina, Siletsky, Konstantinov.This revised version was published online in April 2005 with corrections to the post codes.     

蕲蛇酶配以高压氧治疗急性脑梗死疗效分析

目的 探讨治疗急性脑梗死有效的方案。方法 观察发病在４８ｈ内的急性脑梗死患者７８例,随机分为蕲蛇酶配以高压氧治疗组（高压氧组）和蕲蛇酶治疗组（蕲蛇酶组）。结果 治疗前后两组总有效率分别为９４．７％和９２．５％。基本治愈、显著进步百分比是高压氧组高于蕲蛇酶组,但差异不显著（Ｐ〉０．０５）。神经功能缺损程度总分数治疗前后比较,蕲蛇酶配以高压氧组第１个疗程后,差异显著（Ｐ〈０．０５）,第２个疗程后,差异     

Exogenous GA<Subscript>3</Subscript> and flowering induce the conversion of artemisinic acid to artemisinin in <Emphasis Type="Italic">Artemisia annua</Emphasis> plants

The contents of artemisinin and artemisinic acid were monitored in the Artemisia annua plants treated with GA₃ at vegetative and flowering initiation stages. The highest artemisinin content was observed at full bloom. The decrease in artemisinic acid content occurred during the transition from the vegetative stage to the beginning of flowering. Endogenous GA₃ content in the leaves peaked at full bloom. At the vegetative stage, in plants treated with various concentrations of GA₃ , the content of artemisinin increased while that of artemisinic acid decreased. Apparently, the rate-limiting step in artemisinin biosynthesis was from artemisinic acid to artemisinin. The bottleneck of artemisinin biosynthesis was probably unlocked during the flowering or in the vegetative plants treated with GA₃ , which triggered off the conversion of artemisinic acid to artemisinin.From Fiziologiya Rastenii, Vol. 52, No. 1, 2005, pp. 68–73.Original English Text Copyright © 2005 by Zhang, Ye, Liu, Wang, Li.This article was presented by the authors in English.     

A new medium formulation for <Emphasis Type="Italic">in vitro</Emphasis> rooting of carob tree based on leaf macronutrients concentrations

Experiments were performed to optimize the macronutrients concentrations for in vitro rooting of Ceratonia siliqua micropropagated shoots. Several dilutions of Murashige and Skoog (MS) medium were tested: full-strength MS, half-strength MS ( MS), and MS + full N. The frequency of in vitro rooting was enhanced when the MS was used (50 % rooted shoots). Mature leaves from 20 – 30 year-old carob trees and from 2 year-old micropropagated plants were collected and the concentrations of macronutrients (N, P, K, Ca, Mg) assessed. Based on the mineral composition of the leaves a new medium was formulated and compared with the previous ones showing an increment of the rooting frequency to 80 %. Moreover, shoots rooted in the new medium did not show symptoms of apical necrosis that occurred in the other tested media.