Modifications of the activities of key enzymes and intracellular levels of cyclic nucleotides, in correlation with the glyogen deposition in a cultured hepatoma cell line.

Glycogen accumulation in growing cultures of ZHC cells (originally derived from the Zajdela ascitic hepatoma) is accompanied by an increase in glycogen synthetase (E.C. 2.4.1.11) and phosphorylase (E.C. 2.4.1.1) activities. Essentially the synthetase b and the phosphorylase a are involved in this process. The glycogen accumulation in ZHC cells us preceeded by a noticeable peak of cAMP, whereas cGMP rises early after replating and then decreases simultaneously with the growth rate. The present results suggest that these cultured hepatoma cells undergo throughout every passage an induction process involved in glycogen synthesis storage. Since the original ascites cells growing in vivo (which lack glycogen) and the cultured ZHC cells exhibit similar glycogen synthetase and phosphorylase activities, the resurgence of the glycogenic function (Staedel and Beck, 1978) in the in vitro cultureed cells does not seem related to a change in these two enzymes. By contrast, the high cyclic nucleotide levels in the cultured cells, as compared to those in the ascites cells, offer a possible explanation.     

SO2-induced decrease in photosynthetic activity in two barley cultivars. Evidence against specific damage at the protein-pigment complex level

This research investigated the SO₂-induced effects on photosynthetic apparatus in two barley (Hordeum vulgare L.) cultivars (cv), Panda and Express. Following a chronic treatment with SO₂ (80 ppb, 75 d) neither cv showed any visible signs of injury or chlorosis on leaf surfaces, while a significant reduction in A_max and G_w was detected in both cvs, although to different extents. Thylakoids of SO₂-treated plants showed a decrease in the electron transport activity at the whole chain, photosystem II (PSII) and photosystem I (PSI) level in both cvs. The high performance liquid chromatography (HPLC) analysis of leaf pigments revealed a significant decrease in both cvs, more pronounced in Panda than in Express. Deriphat-polyacrilamide gel electrophoresis (Deriphat-PAGE) and two dimensional (2-D) electrophoretic analyses of the pigment-protein complexes did not show differences in SO₂-treated samples of either cv. HPLC analysis of the green bands also showed no differences in the pigment content of fumigated samples of either cv, except for a decrease in β-carotene content and xanthophyll cycle pigment (VAZ) content respectively at band 1 (PSI) and band 5 (minor light-harvesting polypeptides of PSII) level in cv Panda, where the de-epoxidation index (DEP) significantly increased, while in Express, an increase in VAZ content and DEP value of band 5 was observed. These results suggest that the decrease in the photosynthetic activity can be ascribed, in addition to stomata closure induced by SO₂, to a generalised, rather than specific, pollutant effect on photosynthetic apparatus, which could be interpreted as an adaptation to the adverse environment.     

Immunoaffinity reactors for prion protein qualitative analysis

The cellular prion protein (PrPc) represents the substrate for generation of conformational aberrant PrP isoforms which occur in human and animal prion diseases. The published two-dimensional maps of human PrPc show a vast microheterogeneity of this glycoprotein. The main goal of this project was to develop a highly specific immunoaffinity reactor for qualitative analysis of PrP cellular isoforms isolated from brain homogenate, cerebrospinal fluid and other tissues. New techniques for affinity proteomics, carriers and immobilization chemistry were applied. The choice of matrix (chemical and magnetic properties, particle size and distribution, porosity) was the key factor that influenced the quality of the reactor and the nature of final applications. Mouse anti-prion IgGs directed to N-terminal and C-terminal epitopes (residues 23-40 and 147-165) were grafted in different manners to magnetic micro- and nanoparticles particularly developed for micro-CHIP application. High operational and storage stability of affinity reactors with minimized nonspecific absorption were achieved. The quality of the immunoreactors was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by immunoblotting and by two-dimensional electrophoresis.     

Competitive-level differences in Yo-Yo intermittent recovery and twelve minute run test performance in soccer referees

The aim of this study was to examine yo-yo intermittent recovery test (Yo-Yo test) and 12-minute run test (12MRT) performances in experienced soccer referees of different competitive levels. Three groups (n = 14 each) of experienced Italian soccer referees officiating in the first (series AB, top-level), third (series C, medium-level), and fourth (series D, low-level) division, were randomly submitted to the 12MRT and the Yo-Yo test during 2 testing sessions, 48-hours apart. 12MRT performances were 3,000 +/- 112 m; 2,894 +/- 99 m; and 2,896 +/- 171 m for top-level, medium-level and low-level referees, respectively (p > 0.05). In the Yo-Yo test, the top-level, medium-level, and low-level referees covered 1,874 +/- 431 m; 1,360 +/- 172 m; and 1,272 +/- 215 m, respectively. The test performances of top-level referees in the Yo-Yo test was significantly different from those scored by medium-level and low-level referees (p < 0.05). After the Yo-Yo test, blood lactate concentrations (BLC) were higher in the medium-level and low-level referees compared with the top-level referees (p < 0.05). The results of the present study show that the Yo-Yo test and not the 12MRT can discriminate endurance performance in experienced elite level soccer referees. With respect to its discriminative and match performance validity, the Yo-Yo test may be considered a relevant field test to assess endurance preparedness for experienced soccer referees and a useful tool in talent selection.     

Age-related effects on fitness performance in elite-level soccer referees

Elite soccer referees usually achieve the peak of their officiating careers at an average age that is considerably older than that observed in competitive matched soccer players. As ageing has been reported to negatively affect physical performance, the aim of this study was to investigate the effect of ageing on fitness performance in elite-level soccer referees. Thirty-six elite-level soccer referees were grouped into young (Y, n = 12), average (A, n = 14), and old (O, n = 10) groups, according to their age and observed for field test performance (countermovement jump [CMJ], 50-m and 200-m sprints, and 12-minute running for distance). Results showed a significant age effect on CMJ (r = -0.52, p < 0.001), 200 m (r = 0.51, p < 0.001), and 12-minute time trial running (r = -0.52, p < 0.001). Y jumped higher than A and O groups (p < 0.05) and were faster than O over 200 m (p < 0.05). No group effect was observed for 12-minute run and 50-m performance (p > 0.05), respectively. Sixteen of the 36 referees were further examined for selected physiological variables and grouped into 2 equal (n = 8) age groups (young and old, Y1 and O1, respectively). Vo(2)max was higher in Y1 (p < 0.05), but O1 attained performances similar to Y1 running at selected blood lactate speeds (4 mmol L(-1), p > 0.05). Although older referees demonstrated acceptable fitness levels, younger officials should ensure that they develop appropriate levels of aerobic and anaerobic fitness to be able to match the demands placed upon them while refereeing throughout their careers. To promote this, fitness test standards should be age related.     

Aromatic and aliphatic mono- and bis-nitroxides: a study on their radical scavenging abilities

Nitroxide radicals are an emerging class of interesting compounds with versatile antioxidant and radioprotective properties. All literature studies have so far concentrated on compounds bearing only one nitroxide function. Here, we now investigate and compare the radical scavenging behaviour and antioxidant activity of aromatic indolinonic and aliphatic piperidine bis-nitroxides, i.e compounds bearing two nitroxide functions. Their corresponding mono-derivatives were also studied for comparison. Radical scavenging activity was investigated using EPR and UV-Vis spectroscopy by following spectral changes in acetonitrile of the nitroxides in the presence of alkyl and peroxyl radicals generated, respectively, under anoxic or aerobic conditions from thermal decomposition of AMVN [2,2'-azobis(2,4-di-methylvaleronitrile)]. Antioxidant activity of the nitroxides was evaluated by monitoring conjugated dienes (CD) formation during methyl linoleate micelles peroxidation and by measuring carbonyl content in oxidized bovine serum albumin (BSA). The results show that: (a) each nitroxide moiety in bis-nitroxides scavenges radicals independent of each other; (b) aliphatic nitroxides do not scavenge peroxyl radicals, at least under the experimental conditions used here, whereas indolinonic aromatic ones do: their stoichiometric number is 1.14 and 2.17, respectively, for mono- and bis-derivatives; (c) bis-nitroxides are roughly twice more efficient at inhibiting lipid peroxidation compared to their corresponding mono-derivatives. Although this study provides only comparative information on the relative radical-scavenging abilities of mono- and bis-nitroxides, it helps in understanding further the interesting reactivity of these compounds especially with regards to peroxyl radicals where many controversies in the literature exist.     

Age and sex differences in human skeletal muscle: Role of reactive oxygen species

Previous studies, conducted on experimental animals, have indicated that reactive oxygen species (ROS) are involved in the aging process. The objective of this work was to study the relationship between oxidative damage and human skeletal muscle aging, measuring the activity of the main antioxidant enzymes superoxide dismutase (total and MnSOD), glutathione peroxidase (GPx) and catalase in the skeletal muscle of men and women in the age groups: young (17–40 years), adult (41–65 years) and aged (66–91 years). We also measured glutathione and glutathione disulfide (GSH and GSSG) levels and the redox index; lipid peroxidation and protein carbonyl content. Total SOD activity was lower in the 66–91 year-old vs. the 17–40 year-old men; MnSOD activity was significantly greater in 66–91 year-old vs. 17–40 year-old women. GPx activity remained unchanged. The activity of catalase was lower in adults than in young men but higher in the aged. We observed no changes in GSH levels and significantly higher GSSG levels only in aged men vs. adult men, and a significant decrease in aged women vs. aged men. The protein carbonyl content increased significantly in the 41–65 and 66–91 year-old vs. the 17–40 year-old men. Finally, young women have lower lipid peroxidation levels than young men. Significantly higher lipid peroxidation levels were observed in aged men vs. both young and adult men, and the same trend was noticed for women. We conclude that oxidative damage may play a crucial role in the decline of functional activity in human skeletal muscle with normal aging in both sexes; and that men appear to be more subject to oxidative stress than women.     

Estimation of oxygen uptake from heart rate and ratings of perceived exertion in young soccer players

The objective of this study was to estimate the oxygen uptake (&OV0312;O2) in elite youth soccer players using measures of heart rate (HR) and ratings of perceived exertion (RPEs). Forty-six regional-level male youth soccer players (～13 years) participated in 2 VO(2)max tests. Data for HR, RPE, and VO(2) were simultaneously recorded during the VO(2)max tests with incremental running speed. Regression equations were derived from the first VO(2)max test. Two weeks later, all players performed the same VO(2)max test to validate the developed regression equations. There were no significant differences between the estimated values in the first test and actual values in the second test. During the continuous endurance exercise, the combination of percentage of maximal HR (%HRmax) and RPE measures gave similar estimation of %VO(2)max (R = 83%) in comparison to %HRmax alone (R = 81%). However, the estimation of VO(2) using combined %HRmax and RPE was not satisfactory (R = 45-46%). Therefore, the use of %HRmax (without RPE) to estimate %VO(2)max could be a useful tool in young soccer players during field-based continuous endurance testing and training. Specifically, coaches can use the %HRmax to quantify internal loads (%VO(2)max) and subsequently implement continuous endurance training at appropriate intensities. Furthermore, it seems that RPE is more useful as a measure of internal load during noncontinuous (e.g., intermittent and sprint) exercises but not to estimate %VO(2)max during continuous aerobic exercise (R = 59%).     

Antioxidant activity elicited by low dose of caffeine attenuates pentylenetetrazol-induced seizures and oxidative damage in rats

Although caffeine supplementation has a beneficial effect on people with neurological disorders, its implications for oxidative damage related to seizures are not well documented. Thus the aim of this study was to investigate the effects of two weeks caffeine supplementation (6 mg/kg; p.o.) on seizures and neurochemical alterations induced by pentylenetetrazol (PTZ 60 mg/kg i.p.). Statistical analyses showed that long-term rather than single dose caffeine administration decreased the duration of PTZ-induced seizures in adult male Wistar rats as recorded by cortical electroencephalographic (EEG) and behavioral analysis. The quantification of EEG recordings also revealed that caffeine supplementation protected against a wave increase induced by PTZ. Neurochemical analyses revealed that caffeine supplementation increased glutathione (GSH) content per se and protected against the increase in the levels of thiobarbituric acid reactive substances (TBARS) and oxidized diclorofluoresceine diacetate (DCFH-DA). Also, caffeine prevent the decrease in GSH content and Na⁺, K⁺-ATPase activity induced by PTZ. Our data also showed that the infusion of L-buthionine sulfoximine (BSO; 3.2 μmol/site i.c.v), an inhibitor of GSH synthesis, two days before injecting PTZ reversed the anticonvulsant effect caused by caffeine. BSO infusion also decreased GSH content and Na⁺, K⁺-ATPase activity. However, it increased DCFH-DA oxidation and TBARS per se and reversed the protective effect of caffeine. Results presented in this paper support the neuroprotective effects of low long-term caffeine exposure to epileptic damage and suggest that the increase in the cerebral GSH content caused by caffeine supplementation may provide a new therapeutic approach to the control of seizure.