The aim was to determine whether increased oxidative stress during the adaptation to chronic intermittent hypoxia (CIH) plays a role in the induction of improved cardiac ischemic tolerance. Adult male Wistar rats were exposed to CIH in a hypobaric chamber (7,000 m, 8 h/day, 5 days/wk, 24-30 exposures). Half of the animals received antioxidant N-acetylcysteine (NAC; 100 mg/kg) daily before the exposure; the remaining rats received saline. Control rats were kept under normoxia and treated in a corresponding manner. One day after the last exposure (and/or NAC injection), anesthetized animals were subject to 20 min of coronary artery occlusion and 3 h of reperfusion for determination of infarct size. In parallel subgroups, biochemical analyses of the left ventricular myocardium were performed. Adaptation to CIH reduced infarct size from 56.7 +/- 4.5% of the area at risk in the normoxic controls to 27.7 +/- 4.9%. NAC treatment decreased the infarct size in the controls to 42.0 +/- 3.4%, but it abolished the protection provided by CIH (to 41.1 +/- 4.9%). CIH decreased the reduced-to-oxidized glutathione ratio and increased the relative amount of PKC isoform-delta in the particulate fraction; NAC prevented these effects. The expression of PKC-epsilon was decreased by CIH and not affected by NAC. Activities of superoxide dismutase, catalase, and glutathione peroxidase were affected by neither CIH nor NAC treatment. It is concluded that oxidative stress associated with CIH plays a role in the development of increased cardiac ischemic tolerance. The infarct size-limiting mechanism of CIH seems to involve the PKC-delta-dependent pathway but apparently not the increased capacity of major antioxidant enzymes.

Transgenic plants with genetically increased or decreased levels of cytokinins were used to investigate the effect of cytokinin level on the production of ethylene, a plant hormone with suggested role in senescence, and the production of nitric oxide, potentially important signalling and regulatory molecule. The production of these gases was followed during the course of leaf development and senescence. The production of ethylene and nitric oxide is under genetic control of genes other than those involved in regulation of senescence. The difference in basic ethylene and NO levels in different tobacco cultivars was higher than their changes in senescence. The results of this study did not indicate a direct link between ethylene production and cytokinin levels. However, there was a decreased production of NO in senescent leaves. Low cytokinins level was associated with increased NO production during leaf development. Protein nitrotyrosine proved to be a better indicator of the reactive nitrogen species than measuring of the NO production. Higher nitrotyrosine concentrations were found in insoluble proteins than in the soluble ones, pointing to membrane proteins as the primary targets of the reactive nitrogen species. In plants with elevated cytokinin levels the content of nitrated proteins decreased both in soluble and insoluble fractions. This finding indicates an antioxidative function of cytokinins against reactive nitrogen species.

During ageing of bean (Phaseolus vulgaris L.) cotyledons in plants with modified life span the time-course of four cytokinins, ethylene, and the end products of free radical attack, lipofuscin-like pigments (LFP), were studied. UV irradiation shortened cotyledon life span, while epicotyl decapitation prolonged it. In controls, LFP increased at the senescence onset but at the end of life span it returned to the initial level. Ethylene increased more than 3-fold at the time of abscission. The content of individual cytokinins (zeatin, zeatin riboside, isopentenyl adenine, isopentenyl adenine riboside) varied differently during ageing but they did not decreased in any case under level observed in young cotyledons at the time of abscission. UV irradiation resulted in 14-fold increase in LFP concentration at the end. Ethylene increased 8-fold 2 h after irradiation. Individual cytokinins increased after UV irradiation to a different extent and time-course, nevertheless cotyledon life span was shortened. Decapitation induced LFP decrease. On day 13, LFP abruptly increased and than decreased and stayed lowered until abscission. Ethylene was maximum on day 24, at the time of abscission, it was above 200 % of control. Decapitation produced transient decrease in some cytokinins namely zeatin and isopentenyl adenine riboside.

Ageing of plant organs is accompanied by an increased production of free radicals what results in membrane lipid peroxidation. Non-polar aldehydes originating from this process interact with the cellular material to form the fluorescent end-products, lipofuscin-like pigments (LFP). Their formation was studied both qualitatively and quantitatively in ageing of bean cotyledons. The concentration of lipofuscin-like pigments increased 9-fold in 14-d-old (senescent) cotyledons in relation to 8-d-old (young) cotyledons. HPLC fractionation patterns indicate changes in their composition during ageing. The LFP increase in old cotyledons was accompanied by elevated levels of non-polar aldehydes that increased during ageing to 167 %. The composition of aldehydes was studied by mass spectrometry. The most abundant fraction in both young and old cotyledon was represented by C12 aldehydes, which comprised both saturated and unsaturated species. We have observed differences in abundances of individual aldehydes between the young and the old cotyledons that might explain the differences in the composition of lipofuscin-like pigments. These results support the involvement of free radicals in plant ageing; however, it is suggested that plant aldehydic products of lipid peroxidation differ from those found in animals.

An effect of cyclosporin A on lipid peroxidation in isolated rat hepatocytes was tested. A significant increase in lipid peroxidation marker (the concentration of lipofuscin-like pigments) was observed in samples incubated with cyclosporin A in comparison with the control. When hepatoprotective flavonoid silybin was added, the production of lipofuscin-like pigments decreased significantly. This result indicates a potential positive role of silybin in lowering of cyclosporin A side effects associated with the production of reactive oxygen species and plasma membrane damage.