Enhanced chilling tolerance at the booting stage in rice by transgenic overexpression of the ascorbate peroxidase gene, OsAPXa

Low temperatures during the booting stage reduce rice yields by causing cold-induced male sterility. To determine whether antioxidant capacity affects the ability of rice plants to withstand chilling at the booting stage, we produced transgenic rice plants that overexpress OsAPXa and have increased APX activity. The effect of increased APX activity on the levels of H₂O₂ and lipid peroxidation were determined by measuring H₂O₂ levels and malondialdehyde (MDA) contents in spikelets during cold treatments at the booting stage. The levels of H₂O₂ and the MDA content increased by 1.5-fold and twofold, respectively in WT plants subjected to a 12°C treatment for 6 days. In contrast, transgenic lines showed small changes in H₂O₂ levels and MDA content under cold stress, and H₂O₂ levels and MDA content were significantly lower than in WT plants. APX activity showed negative correlations with levels of H₂O₂ and MDA content, which increased during cold treatment. Cold tolerance at the booting stage in transgenic lines and WT plants was evaluated. Spikelet fertility was significantly higher in transgenic lines than in WT plants after a 12°C treatment for 6 days. These results indicate that higher APX activity enhances H₂O₂-scavenging capacity and protects spikelets from lipid peroxidation, thereby increasing spikelet fertility under cold stress.     

Cytochrome P450 oxidase activity and its role in NADPH dependent lipid peroxidation

A comparison is made between microsomal NADPH-dependent H₂O₂ production and malondialdehyde (MDA) formation in rat liver microsomes, obtained from phenobarbital pretreated rats. An increase in H₂O₂ formation was observed during NADPH-dependent disposition (10 min) of 100 μM diazepam (33%) and 2 mM hexobarbital (69%). In contrast orphenadrine (100 μM) and its mono-N-demethylated metabolite tofenacine (100 μM) decreased the H₂O₂ formation (35% and 55%, respectively). However, all these substrates were found to inhibit NADPH-dependent lipid peroxidation (60 min), estimated by measuring MDA formation, to various extents. These data strongly suggest that the oxidase activity of cytochrome P450 (H₂O₂ production) is not involved in a rate-limiting step in NADPH-dependent lipid peroxidation.     

Maternal selenium deficiency increases hydrogen peroxide and total lipid peroxides in porcine fetal liver

To investigate the role of selenium (Se) in the developing porcine fetus, prepubertal gilts (n=42) were randomly assigned to either Se-adequate (0.39 ppm Se) or Se-deficient (0.05 ppm Se) gestation diets 6 wk prior to breeding. Maternal and fetal liver was collected at d 30, 45, 70, 90, and 114 of pregnancy. Concentrations of Se in maternal liver decreased during gestation in gilts fed the low-Se diet. The activity of cellular glutathione peroxidase (GPx) was decreased at d 30 and 45 of gestation in liver of gilts fed the low-Se diet. Concentrations of malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) were greater in liver homogenates from gilts fed the low-Se diet. Within the fetuses, liver Se decreased in those fetuses of gilts fed the low-Se diet. Although the activity of GPx in fetal liver was not affected by the maternal diet, concentrations of H₂O₂ and MDA in fetal liver were greater in fetuses from gilts fed the low-Se diet. Maternal liver GPx activity was approx 12-fold greater than fetal liver GPx activity regardless of dietary treatment. These results indicate that maternal dietary Se intake affects fetal liver Se concentration and feeding a low-Se diet during gestation increases oxidative stress to the fetus, as measured by fetal liver H₂O₂ and MDA.     

Effect of hydrogen peroxide on contractility and citrate synthase activity of the rabbit urinary bladder in the presence and absence of resveratrol and a whole-grape suspension

One etiology related directly to obstructive urinary bladder dysfunction is ischemia/reperfusion resulting in significant oxidative stress to the bladder. Grapes, a natural source of antioxidants, have been proven effective in preventing obstructive and ischemic bladder dysfunction. Many investigators believe that resveratrol is the primary active antioxidant ingredient in grapes. We compared the ability of a whole-grape suspension with pure resveratrol in their ability to protect the bladder from in vitro oxidative stress mediated by hydrogen peroxide (H₂O₂). Four male rabbit bladders were used. Two strips from each bladder were incubated in the presence of 1 mg/mL grape suspension for 30 min, another two strips were incubated in the presence of 1 mg/mL resveratrol solution, and the last two strips were incubated in the presence of 1 mg/mL sucrose/and fructose as controls. The rest of the bladder was separated into muscle and mucosa, frozen and stored for biochemical evaluation. (1) Chemically, resveratrol has about 20 times the antioxidant capacity of the grape suspension. (2) The grape suspension had significant protective effects when the rate of tension was quantitated at all concentrations of H₂O₂, while the resveratrol had no effect. (3) Citrate synthase activities of the muscle and mucosa were significantly protected by the grape suspension but not by resveratrol. These data demonstrate that the grape suspension protects the mitochondria to a significantly greater degree than resveratrol, which suggests that the antioxidant activities are due to the combination of active components found in the grape suspension and not just resveratrol.     

Contraction of guinea pig gail bladder strips by leukotrienes and other agonists

In the presence of indomethacin, Leukotriene C₄ (LTC₄), LTD₄ and LTE₄ were shown to be contractile agents on guinea pig gall bladder strips. The respective pD₂ values for LTC₄, LTD₄ ad LTE₄ were 9.1, 9.1 and 7.7. The contractile effects of LTD₄ were not mediated through the generation of cyclooxygenase products and were antagonized by the SRS-A antagonist FPL-55712. The effects of PGE₁, PGF_2α, the endoperoxide analogue U44069 and histamine on gall bladder strips were also examined. All these agents caused dose-related contractions but were considerably less potent than the leukotrienes. Leukotrienes are therefore potent contractile agents on the guinea pig gall bladder and may contribute to gall bladder contractions or spasms .     

Myocardial glutathione metabolic status in fat-fed rabbits

Short-term fat feeding could exert adverse cardiac effects by altering myocardial glutathione-related antioxidant defenses. We have here assessed total glutathione (TG), the activities of glutathione reductase (GSSG-Red), γ-glutamylcysteine synthetase (γ-GCS), γ-glutamyl transpeptidase (γ-GT) and glutathione peroxidase (GSH-Px), fluorescent damage products of lipid peroxidation (FDPL), thiobarbituric acid-reactive substances (TBARS), H₂O₂, and ATP in the aerobically perfused hearts of control rabbits and of rabbits fed a fat-enriched diet for 18 days. Such biochemical parameters, myocardial hemodynamics and infarct size were assessed in the perfused hearts of other control and fat-fed rabbits subjected to 60 min global ischemia plus 30 min reperfusion. Compared to controls, a reduced activity of GSSG-Red and γ-GT associated with decreased TG content was detected in the aerobically perfused hearts of fat-fed rabbits, which also showed insignificant γ-GCS activation, GSH-Px depressed activity, FDPL, TBARS and H₂O₂ burden, and unaltered ATP content. Ischemia–reperfusion decreased the myocardial levels of TG, ATP, and γ-GCS activity and augmented those of FDPL, TBARS, and H₂O₂ especially in the fat-fed rabbits, without significant changes in myocardial GSSG-Red, γ-GT, and GSH-Px activities. Ischemia–reperfusion induced greater hemodynamic dysfunction and infarct size in the hearts of fat-fed rabbits than in those of controls. Thus, short-term fat feeding and hyperlipidemia alter glutathione metabolic status of the rabbit myocardium, inducing a GSSG-Red- and γ-GT-related decrement of myocardial glutathione content, which, together with GSH-Px dysfunction, may favor tissue oxidative stress and render the myocardium more susceptible to ischemia–reperfusion injury.     

Thromboxane A2 antagonist inhibits leukotriene D4-induced smooth muscle contraction in guinea-pig lung parenchyma, but not in trachea

Although the bronchoconstriction induced by leukotriene D₄ (LTD₄) has been reported to be partly mediated by thromboxane A₂ (TXA₂) in the guinea-pig airway, it is not known which part of the airway is susceptible to TXA₂. In order to determine the role of TXA₂ in the central and peripheral airways, we compared the effect of a TXA₂ antagonist on tracheal strips to its effect on parenchymal strips of guinea-pigs. Tracheal and parenchymal strips were mounted in a 3.5 ml organ bath filled with Krebs-Henseleit solution aerated with 95% O₂, 5% CO₂ and kept at 37°C. After equilibration for 60 min in Krebs solution, the strip was contracted by exposure to 10⁻⁵ M of acetylcholine (ACh). Sixty minutes after ACh was eliminated, the concentration-response curve to LTD₄ (10⁻⁹ M–10⁻⁷ M) was obtained, and the LTD₄-induced contractions were expressed as the percent of the contraction evoked by 10⁻⁵ M of ACh. We measured the contractile response to LTD₄ in the presence or absence of the TXA₂ antagonist, BAY u3405 (10⁻⁸ M–10⁻⁶ M). In the tracheal strips, BAY u3405 had no effect on the LTD₄-induced contraction. However, in parenchymal strips, BAY u3405 significantly suppressed the contractile response to LTD₄. These results suggest that in the central airway LTD₄ contracts smooth muscle directly, but that in the peripheral airway LTD₄ induces smooth muscle contraction both directly and indirectly, via TXA₂.     

Increase of pro-oxidants with no evidence of lipid peroxidation in exhaled breath condensate after a 10-km race in non-athletes

It is a well-established fact that exercise increases pro-oxidants and favors oxidative stress; however, this phenomenon has been poorly studied in human lungs. Pro-oxidative generation (H₂O₂, NO₂ ^?), lipid peroxidation markers (MDA), and inflammation (pH) in exhaled breath condensate (EBC) have been determined through data from 10 active subjects who ran 10 km; samples were obtained immediately before, at 20, and at 80 min post-exertion. In EBC, the concentration of H₂O₂ at 80 min post-exertion was increased. NO₂ ^? concentration showed a tendency to increase at 80 min post-exertion, with no variations in MDA and pH. No variations of NO₂ ^? were found in plasma, while there was an increase of NO₂ ^? at 80 min post-exertion in the relation between EBC and plasma. NO₂ ^? in EBC did not correlate to plasmatic NO₂ ^?, while it did correlate directly with H₂O₂ in EBC, suggesting a localized origin for the exercise-related NO₂ ^? increase in EBC. MDA in plasma did not increase nor correlate with MDA in EBC. In conclusion, high-intensity exercise increases lung-originated pro-oxidants in non-athlete subjects with no evidence of early lipid peroxidation and changes in the pH value in EBC.     

Exogenous proline alleviates the effects of H2O2-induced oxidative stress in wild almond species

The effect of proline on the antioxidant system in the leaves of eight species of wild almond (Prunus spp.) exposed to H₂O₂-mediated oxidative stress was studied. The levels of endogenous proline (Pro) and hydrogen peroxide, and the activities of total superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), and guaiacol peroxidase (POD) were measured. The degradation of chlorophyll but not carotenoids occurred in leaves in the solution of 5 mM H₂O₂. An increase in membrane lipid peroxidation was observed in H₂O₂ treatment, as assessed by MDA level and percentage of membrane electrolyte leakage (EL). Significant increases in total SOD and CAT activities, as well as decreases in APX and POD activities, were detected in H₂O₂-treated leaves. The three SOD isoforms showed different behavior, as Mn-SOD activity was enhanced by H₂O₂, whereas Fe-SOD and Cu/Zn-SOD activities were inhibited. In addition, Pro accumulation up to 0.1 ??mol/g fr wt, accompanied by significant decreases in ascorbate and glutathione levels, was observed in H₂O₂-treated leaves. After two different treatments with 10 mM Pro + 5 mM H₂O₂, total SOD and CAT activities were similar to the levels in control plants, while POD and APX activities were higher if compared to the leaves exposed only to H₂O₂. Pro + H₂O₂ treatments also caused a strong reduction in the cellular H₂O₂ and MDA contents and EL. The results showed that Pro could have a key role in protecting against oxidative stress injury of wild almond species by decreasing membrane oxidative damage.     

Selective Alkylation of Rat Urinary Bladder Muscarinic Receptors with 4-DAMP Mustard Reveals a Contractile Function for the M2 Muscarinic Receptor

Abstract

Our previous data indicate that M₃ muscarinic receptors mediate carbachol induced bladder contractions. The data presented here were obtained by selective alkylation of M₃ receptors with 4-DAMP mustard and suggest that the M₂ receptor subtype may be involved in inhibition of β-adrenergic receptor induced relaxation, therefore, allowing recontraction. Alkylation resulted in 85% of M₃ receptors and 65% of M₂ receptors unable to bind radioligand as demonstrated by subtype selective immunoprecipitation. Rat bladder strips subjected to our alkylation procedure contracted submaximally, and direct carbachol contractions were inhibited by antagonists with affinities consistent with M₃ receptor mediated contraction. In contrast, the affinities of antagonists for inhibition of carbachol induced recontractions following isoproterenol stimulated relaxation in the presence of 90 mM KCI, indicated a contractile function for the M₂ receptor that was not observed in control strips. In conclusion, these studies demonstrate a possible role for the M₂ subtype in bladder smooth muscle contraction.     

Histamine H2-receptors in guinea pig peripheral airway smooth muscle.

The presence and function of histamine H₂-receptors in guinea pig lung was studied using lung strips as an in vitro model of peripheral airway smooth muscle. The lung strips were incubated in Krebs-Henseleit solution in the absence or presence of specific antagonists for 20 min prior to the addition of either histamine or dimaprit added in a half-log cumulative fashion. Changes in isometric tension were recorded. Histamine at low concentrations (10^?7?10^?6M) caused a slight relaxation which was potentiated by the histamine H₁-antagonist chlorpheniramine (10^?7 or 10^?6M) and abolished by the histamine H₂-antagonist metiamide (10^?4M). Higher concentrations of histamine produced a dose-related contraction which was antagonized competitively by chlorpheniramine or potentiated by metiamide. Dimaprit, a histamine H₂-agonist, produced only a relaxant response over the concentration range of 10^?7 ? 10^?3M. This relaxation was reduced by metiamide but not by the beta adrenergic antagonist propranolol. These results indicate the presence of both histamine H₂ and H₁-receptors in guinea pig peripheral airway smooth muscle which mediate the relaxant and contractile effects of histamine respectively.     

Hydrogen peroxide pretreatment of roots enhanced oxidative stress response of tomato under cold stress

In the view of physiological role of H₂O₂, we investigated whether exogenous H₂O₂ application would affect short-term cold response of tomato and induce acclimation. Pretreatments were performed by immersing roots into 1 mM H₂O₂ solution for 1 h when transferring seedlings from seedling substrate to soil (acclimated group). Cold stress (3 °C for 16 h) caused significant reduction in relative water content (RWC) of control and non-acclimated (distilled water treated) groups when compared with unstressed plants. H₂O₂ promoted maintenance of relatively higher RWC under stress. Anthocyanin level in leaves of acclimated plants under cold stress was significantly higher than that of unstressed control and non-acclimated plants. Malondialdehyde (MDA) levels demonstrated low temperature induced oxidative damage to control and non-acclimated plants. MDA remained around unstressed conditions in acclimated plants, which demonstrate that H₂O₂ acclimation protected tissues against cold induced lipid peroxidation. H₂O₂ acclimation caused proline accumulation in roots under cold stress. Ascorbate peroxidase (APX) activity in roots of cold stressed and unstressed H₂O₂ acclimated plants increased when compared with control and non-acclimated plants, with highest increase in roots of acclimated plants under cold stress. CAT levels in roots of acclimated plants also increased, whereas levels remained unchanged in unstressed plants. Endogenous H₂O₂ levels significantly increased in roots of control and non-acclimated plants under cold stress. On the other hand, H₂O₂ content in roots of acclimated plants was significantly lower than control and non-acclimated plants under cold stress. The results presented here demonstrated that H₂O₂ significantly enhanced oxidative stress response by elevating the antioxidant status of tomato.     

Chloroplastic NADPH oxidase-like activity-mediated perpetual hydrogen peroxide generation in the chloroplast induces apoptotic-like death of <Emphasis Type="Italic">Brassica napus</Emphasis> leaf protoplasts

Despite extensive research over the past years, regeneration from protoplasts has been observed in only a limited number of plant species. Protoplasts undergo complex metabolic modification during their isolation. The isolation of protoplasts induces reactive oxygen species (ROS) generation in Brassica napus leaf protoplasts. The present study was conducted to provide new insight into the mechanism of ROS generation in B. napus leaf protoplasts. In vivo localization of H₂O₂ and enzymes involved in H₂O₂ generation and detoxification, molecular antioxidant-ascorbate and its redox state and lipid peroxidation were investigated in the leaf and isolated protoplasts. Incubating leaf strips in the macerating enzyme (ME) for different duration (3, 6, and 12 h) induced accumulation of H₂O₂ and malondialdehyde (lipid peroxidation, an index of membrane damage) in protoplasts. The level of H₂O₂ was highest just after protoplast isolation and subsequently decreased during culture. Superoxide generating NADPH oxidase (NOX)-like activity was enhanced, whereas superoxide dismutase (SOD) and ascorbate peroxidase (APX) decreased in the protoplasts compared to leaves. Diaminobenzidine peroxidase (DAB-POD) activity was also lower in the protoplasts compared to leaves. Total ascorbate content, ascorbate to dehydroascorbate ratio (redox state), were enhanced in the protoplasts compared to leaves. Higher activity of NOX-like enzyme and weakening in the activity of antioxidant enzymes (SOD, APX, and DAB-POD) in protoplasts resulted in excessive accumulation of H₂O₂ in chloroplasts of protoplasts. Chloroplastic NADPH oxidase-like activity mediated perpetual H₂O₂ generation probably induced apoptotic-like cell death of B. napus leaf protoplasts as indicated by parallel DNA laddering and decreased mitochondrial membrane potential.     

Effect of partial outlet obstruction on nitrotyrosine content and distribution within the rabbit bladder

Purpose: Evidence indicates that free radicals are etiological factors in obstructive bladder disease. However, it is not clear which species of reactive oxygen or nitrogen species mediate the damage. The current studies were designed to determine if partial outlet obstruction in rabbits results in the generation of nitrotyrosine (NT). Materials and methods: Sixteen rabbits were separated into four groups of four. The rabbits in groups 1 and 2 underwent sham operation while rabbits in groups 3 and 4 underwent partial outlet obstruction. The rabbits in groups 1 and 3 were evaluated after 1 week of obstruction and the rabbits in groups 2 and 4 were evaluated after 2 weeks of obstruction. A separate group of four controls were evaluated simultaneously with the sham and obstructed rabbits. Four rabbits from each group were evaluated after 1 and 2 weeks of obstruction. Four control rabbits were also evaluated. Isolated strips were evaluated for contractile responses and NT content of the mucosa and muscle were quantitated by Western blot analysis. Results: (1) The mucosa contains both 42 and 62 kD proteins exhibiting a strong nitrotyrosine signal; the muscle presents a signal only at 62 kD. (2) The sham operations had no effect on nitrotyrosine distribution or content. (3) The nitrotyrosine of both mucosal proteins and the muscle protein are increased in the 1 week obstructed bladder; whereas, only the 62 kD signal is increased in the two week obstructed bladder mucosa. (4) The contractile response to FS are reduced to a significantly greater degree than the responses to carbachol, KCl, or ATP. Conclusions: These studies clearly demonstrated that partial outlet obstruction in rabbits results in significant increases in nitrotyrosine within the bladder and may contribute to the contractile dysfunctions mediated by partial outlet obstruction. (Mol Cell Biochem 276: 143–148, 2005)     

Interactive effects of salicylic acid and silicon on oxidative damage and antioxidant activity in spinach (<Emphasis Type="Italic">Spinacia oleracea</Emphasis> L. cv. Matador) grown under boron toxicity and salinity

We investigated individual and combined effects of salinity, soil boron (B), silicon (Si) and salicylic acid (SA) on the activities of major antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT and ascorbate peroxidase, APX) and non-enzymatic antioxidants (AA), proline, chlorophyll, anthocyanin, H₂O₂ concentration, stomatal resistance (SR), lipid peroxidation (MDA), membrane permeability (MP), and the uptake of sodium (Na), chloride (Cl), boron and Si of spinach plants. In general, salinity significantly increased H₂O₂ and proline concentrations, antioxidant activity, membrane permeability, lipid peroxidation and SR of the spinach plants, indicating that they were stressed, whereas application of B only increased proline concentration. However, plant fresh weights did not decline with either treatment. The application of Si decreased H₂O₂ and increased the activity of SOD and CAT. The application of SA increased SOD activity. Neither SA nor Si had any effect on the proline concentration, or MP. However, application of Si increased chlorophyll concentration and decreased lipid peroxidation (MDA concentration). Si treatment had no effect on SR. The concentration of B in the tissues, which was strongly increased by B treatment, was decreased by NaCl. As a result of salinity, concentrations of Na⁺ and Cl⁻ ions were increased in the plant tissues, and application of Si slightly increased these concentrations. These results indicate that exogenous Si application increases stress tolerance of spinach, a plant that is naturally reasonably resistant to combined salinity and B toxicity, by the enhancement of antioxidant mechanisms that reduce membrane damage. Exogenous SA has a less obvious effect, although the levels of salinity and boron stress applied were not sufficient in this experiment to reduce plant fresh weight.     

Hydrogen peroxide-induced oxidative stress to the mammalian heart-muscle cell (cardiomyocyte): Lethal peroxidative membrane injury

Oxidative stress induced by hydrogen peroxide (H₂O₂) may contribute to the pathogenesis of ischemic-reperfusion injury in the heart. For the purpose of investigating directly the injury potential of H₂O₂ on heart muscle, a cellular model of H₂O₂-induced myocardial oxidative stress was developed. This model employed primary monolayer cultures of intact, beating neonatal-rat cardiomy-ocytes and discrete concentrations of reagent H₂O₂ in defined, supplement-free culture medium. Cardiomyocytes challenged with H₂O₂ readily metabolized it such that the culture content of H₂O₂ diminished over time, but was not depleted. The consequent H₂O₂-induced oxidative stress caused lethal sarcolemmal disruption (as measured by lactate dehydrogenase release), and cardiomyocyte integrity could be preserved by catalase. During oxidative stress, a spectrum of cellular derangements developed, including membrane phospholipid peroxidation, thiol oxidation, consumption of the major chain-breaking membrane antiperoxidant (α-tocopherol), and ATP loss. No net change in the protein or phospholipid contents of cardiomyocyte membranes accompanied H₂O₂-induced oxidative stress, but an increased turnover of these membrane constituents occurred in response to H₂O₂. Development of lethal cardiomyocyte injury during H₂O₂-induced oxidative stress did not require the presence of H₂O₂ itself; a brief “pulse” exposure of the cardiomyocytes to H₂O₂ was sufficient to incite the pathogenic mechanism leading to cell disruption. Cardiomyocyte disruption was dependent upon an intracellular source of redox-active iron and the iron-dependent transformation of internalized H₂O₂ into products (e.g., the hydroxyl radical) capable of initiating lipid peroxidation, since iron chelators and hydroxyl-radical scavengers were cytoprotective. The accelerated turnover of cardiomyocyte-membrane protein and phospholipid was inhibited by antiperoxidants, suggesting that the turnover reflected molecular repair of oxidized membrane constituents. Likewise, the consumption of α-tocopherol and the oxidation of cellular thiols appeared to be epiphenomena of peroxidation. Antiperoxidant interventions coordinately abolished both H₂O₂-induced lipid peroxidation and sarcolemmal disruption, demonstrating that an intimate pathogenic relationship exists between sarcolemmal peroxidation and lethal compromise of cardiomyocyte integrity in response to H₂O₂-induced oxidative stress. Although sarcolemmal peroxidation was causally related to cardiomyocyte disruption during H₂O₂-induced oxidative stress, a nonperoxidative route of H₂O₂ cytotoxicity was also identified, which was expressed in the complete absence of cardiomyocyte-membrane peroxidation. The latter mode of H₂O₂-induced cardiomyocyte injury involved ATP loss such that membrane peroxidation and cardiomyocyte disruption on the one hand and cellular de-energization on the other could be completely dissociated. The cellular pathophysiology of H₂O₂ as a vectorial signal for cardiomyocyte necrosis that “triggers” irreversible peroxidative disruption of the sarcolemma has implications regarding potential mechanisms of oxidative injury in the postischemic heart.     

Accelerated ageing and subsequent imbibition affect seed viability and the efficiency of antioxidant system in macaw palm seeds

Accelerated ageing is an accurate test indicator of seed vigor and storability that helps to understand the mechanisms of cellular and biochemical deterioration that occur during seed ageing. This study was carried out to elucidate the mechanisms of ageing in macaw palm embryos. Seeds were artificially aged during 4, 8 and 12 days at 45 °C and 100% relative humidity. After ageing, seeds were tested for viability (tetrazolium), electrical conductivity, lipid peroxidation (MDA) and hydrogen peroxide (H₂O₂) content. Part of the aged seeds was imbibed for 8 days and then determined the hydrogen peroxide content and the activity of antioxidant system enzymes (superoxide dismutase, catalase and glutathione reductase). Ageing reduced the embryo viability from 8 days of treatment and increased malondialdehyde content (MDA) and solute leakage. Hence, membrane permeability correlated with both loss of viability and lipid peroxidation. Imbibition after ageing significantly increased H₂O₂ content along with superoxide dismutase activity. Catalase activity was significantly higher than control in embryos aged from 8 days and imbibed, and glutathione reductase activity did not change. Our results suggest that macaw palm seed deterioration during accelerated ageing is closely related to lipid peroxidation, and that enzymatic antioxidant system is not completely efficient in reducing reactive oxygen species after imbibition, a critical phase to germination. Moreover, accelerated ageing test can be used as a reliable model to understand the mechanisms involved in palm seeds deterioration.     

Effect of Oxidative Stress on the Expression of Thin Filament-Associated Proteins in Gastric Smooth Muscle Cells

Thin filament-associated proteins such as calponin, caldesmon, and smoothelin are believed to regulate acto-myosin interaction and thus, muscle contraction. Oxidative stress has been found to affect the normal contractile behavior of smooth muscle and is involved in the pathogenesis of a number of human diseases such as diabetes mellitus, hypertension, and atherosclerosis. However, very little is known about the effect of oxidative stress on the expression of smooth muscle contractile proteins. The aim of the current study is to investigate the effect of oxidative stress on the expression of thin filament-associated proteins in rat gastric smooth muscle. Single smooth muscle cells of the stomach obtained from Sprague–Dawley rats were used. Muscle cells were treated with hydrogen peroxide (H₂O₂) (500 μM) for 30 min or the peroxynitrite donor 3-morpholinosydnonimine (SIN-1) (1 mM) for 90 min to induce oxidative stress. Calponin, caldesmon, and smoothelin expressions were measured via specifically designed enzyme-linked immunosorbent assay. We found that exposure to exogenous H₂O₂ or incubation of dispersed gastric muscle cells with SIN-1 significantly increased the expression of calponin, caldesmon, and smoothelin proteins. In conclusion: oxidative stress increases the expression of thin filament-associated proteins in gastric smooth muscle, suggesting an important role in gastrointestinal motility disorders associated with oxidative stress.     

Nitric oxide retards xanthine oxidase-mediated superoxide anion generation in Phalaenopsis flower: an implication of NO in the senescence and oxidative stress regulation

Senescence is a developmentally regulated and highly ordered sequence of events. Senescence leads to abscission of plant organs and eventually leads to death of a plant or part of it. Present study revealed that Phalaenopsis flower undergo senescence due to over activation of O₂ ^·−generating xanthine oxidase (XO), which consequently increases the concentrations of O₂ ^·− leading to enhanced oxidative damage and disturbed cellular redox environment as indicated by increased lipid peroxidation and DHA/AsA + DHA ratio, respectively. While activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), and non-specific peroxidase (POD) were enhanced in sepals and petals of old flower, activities of catalase (CAT) and glutathione reductase (GR) were decreased. Exogenous application of nitric oxide (NO) retarded H₂O₂-induced senescence of Phalaenopsis flower by downregulating activity of XO and concentrations of O₂ ^·−, H₂O₂ and malondialdehyde (MDA, an index of lipid peroxidation). Exogenous application of NO also downregulated SOD activity and upregulated antioxidant enzymes involved in the detoxification of H₂O₂ (CAT and APX), and in the regulation of redox couples viz, monodehydroascorbate reductase (MDHAR) and GR, together with the modulation in non-protein thiol status and DHA/AsA + DHA ratio.     

Endogenous hydrogen sulfide regulation of myocardial injury induced by isoproterenol

Previous work has shown that the endogenous cystathionine γ-synthase (CSE)/hydrogen sulfide (H₂S) pathway participates in the regulation of cardiac contraction. We hypothesized that the pathway might participate in the pathophysiological regulation of ischemic heart disease. Isoproterenol injection of rat hearts induced a myocardial ischemic injury model, with reduced myocardial and plasma H₂S levels, decreased CSE activity, and upregulated CSE gene expression. Exogenous administration of the H₂S donor NaHS reduced the mortality rate; increased left-ventricular pressure development and left-ventricular-end systolic pressure; and decreased left-ventricular-end diastolic pressure (LVEDP) and subendocardial necrosis, capillary dilatation, leukocytic infiltration, fibroblast swelling, and fibroblastic hyperplasia. As well, production of lipid peroxidation, including myocardial malondialdehyde (MDA), and plasma MDA and conjugated diene, was reduced. Oxidative stress injury is an important mechanism of isoproterenol-induced myocardial injury. In vitro experiments revealed that NaHS might antagonize myocyte MDA production by oxygen-free radicals and that NaHS directly scavenged hydrogen peroxide and superoxide anions. Our results suggest that the endogenous CSE/H₂S pathway contributes to the pathogenesis of isoproterenol-induced myocardial injury. Administration of exogenous H₂S effectively protects myocytes and contractile activity, at least by its direct scavenging of oxygen-free radicals and reducing the accumulation of lipid peroxidations.