Reactive oxygen species accumulation patterns of alfalfa root nodules identified using an optimized method

ROS include superoxide anion radical (O2^-), hydrogen peroxide (H2O2), singlet oxygen (0-0), hydroxyl radical (OH ), and hydroxyl anion (OH-). O2^-, the radical with the highest oxidation compacity to produce H2O2, OH and OH^-, is a byproduct of aerobic respiration in organisms. Rhizobia and their legume hosts are aerobic organisms, and nitrogen-fixing nodules formed on legume roots are developed from their symbiotic in teractions. Therefore, ROS are generated in legume nodules through aerobic respiration by both symbiotic partners.     

Type 2 diabetes caused by reductive redox potential?

<正>Reactive oxygen species arise(ROS)in the mitochondria as byproducts of respiration and oxidase activity and have important roles in many physiological and pathophysiological conditions.The current literature indicate that excessive levels of ROS can cause oxidative stress and that lots of evidences link ROS and oxidative stress to the pathogenesis of type 2 diabetes mellitus(T2DM)and development of complications.Several studies have shown elevated extraand intracellular glucose concentrations result in oxidative stress both in animal models of diabetes and in diabetic patients[1].And ROS can contribute to the development and progression of diabetes and related complications by directly damaging DNA,proteins,and lipids or indirectly activating a number of cellular stress-sensitive pathways to induce damage to tissues such as isletβcells[2].     

Mutual interaction between oxidative stress and endoplasmic reticulum stress in the pathogenesis of diseases specifically focusing on non-alcoholic fatty liver disease

Reactive oxygen species(ROS) are produced during normal physiologic processes with the consumption of oxygen. While ROS play signaling roles, when they are produced in excess beyond normal antioxidative capacity this can cause pathogenic damage to cells. The majority of such oxidation occurs in polyunsaturated fatty acids and sulfhydryl group in proteins, resulting in lipid peroxidation and protein misfolding, respectively. The accumulation of misfolded proteins in the endoplasmic reticulum(ER) is enhanced under conditions of oxidative stress and results in ER stress, which, together, leads to the malfunction of cellular homeostasis. Multiple types of defensive machinery are activated in unfolded protein response under ER stress to resolve this unfavorable situation. ER stress triggers the malfunction of protein secretion and is associated with a variety of pathogenic conditions including defective insulin secretion from pancreatic β-cells and accelerated lipid droplet formation in hepatocytes. Herein we use nonalcoholic fatty liver disease(NAFLD) as an illustration of such pathological liver conditions that result from ER stress in association with oxidative stress. Protecting the ER by eliminating excessive ROS viathe administration of antioxidants or by enhancing lipidmetabolizing capacity via the activation of peroxisome proliferator-activated receptors represent promising therapeutics for NAFLD.     

Reactive oxygen species: Multidimensional regulators of plant adaptation to abiotic stress and development

Reactive oxygen species(ROS) are produced as undesirable by-products of metabolism in various cellular compartments, especially in response to unfavorable environmental conditions,throughout the life cycle of plants. Stressinduced ROS production disrupts normal cellular function and leads to oxidative damage. To cope with excessive ROS, plants are equipped with a sophisticated antioxidative defense system consisting of enzymatic and non-enzymatic components that scavenge ROS or inhibit their har...     

Altered Redox Status in Erythrocytes From Hypertensive Subjects： Effect of （-）Epicatechin

Hypertension is a major problem worldwide. There is much evidence to suggest that reactive oxygen species (ROS) radical may play a role in the development of organ damage associated with cardiovascular disease and hypertension. ( － )Epicatechin, a member of tea catechins belonging to flavonoid group, is known to be a potent anti-oxidant. The study has been undertaken to evaluate the effect of ( － )epicatechin on markers of oxidative stress: reduced glutathione (GSH) and membrane sulfhydryl ( — SH) groups in erythrocytes from hypertensive patients. The effect of ( － )epicatechin was also compared with a known anti-oxidant L-ascorbic acid. The erythrocyte intracellular GSH content and membrane — SH group content were significantly (P<0.01) decreased in hypertensive subjects. In vitro incubation with ( － )epicatechin caused an increase in GSH and — SH content, the effect was more pronounced in hypertensive erythrocytes. Similar results were obtained with L-ascorbic acid. The observed decrease in the level of GSH and — SH groups in hypertension is an indicator of oxidative stress condition. Observation of an increase in red cell GSH content and the protection of membrane — SH group oxidation by ( － )epicatechin in hypertensive subjects is a convincing reason to suggest that high dietary intake of foods rich in catechins may help to reduce oxidative stress and concomitant free radical damage in hypertensive patients.     

Hydrogen Peroxide in Plants： a Versatile Molecule of the Reactive Oxygen Species Network

Plants often face the challenge of severe environmental conditions, which include various biotic and abiotic stresses that exert adverse effects on plant growth and development. During evolution, plants have evolved complex regulatory mechanisms to adapt to various environmental stressors. One of the consequences of stress is an increase in the cellular concentration of reactive oxygen species （ROS）, which are subsequently converted to hydrogen peroxide （H2O2）. Even under normal conditions, higher plants produce ROS during metabolic processes. Excess concentrations of ROS result in oxidative damage to or the apoptotic death of cells. Development of an antioxidant defense system in plants protects them against oxidative stress damage. These ROS and, more particularly, H2O2, play versatile roles in normal plant physiological processes and in resistance to stresses. Recently, H2O2 has been regarded as a signaling molecule and regulator of the expression of some genes in cells. This review describes various aspects of H2O2 function, generation and scavenging, gene regulation and cross-links with other physiological molecules during plant growth, development and resistance responses.     

2-Bromopalmitate attenuates inflammatory pain by maintaining mitochondrial fission/fusion balance and function

Inflammatory pain activates astrocytes and increases inflammatory cytokine release in the spinal cord. Mitochondrial fusion and fission rely on the functions of dynamin-related protein 1 (Drp1) and optic atrophy 1 (OPA1), which are essential for the synaptic transmission and plasticity. In the present study, we aimed to explore the effects of 2-bromopalmitate (2-BP), an inhibitor of protein palmitoylation, on the modulation of pain behavior. Rats were intraplantar injected with complete Freund’s adjuvant (CFA) to establish an inflammatory pain model. In the spinal cord of rats with CFA-induced inflammatory pain, the expression of astrocyte-specific glial fibrillary acidic protein (GFAP) and contents of proinflammatory cytokines IL-1β and TNF-α were increased. Mitochondrial Drp1 was increased, while OPA1 was decreased. Consequently, CFA induced reactive oxygen species (ROS) production and Bcl-2-associated X protein (BAX) expression. The intrathecal administration of 2-BP significantly reversed the pain behaviors of the inflammatory pain in rats. Moreover, 2-BP also reduced the Drp1 expression, elevated the OPA1 expression, and further reduced the GFAP, IL-1β, and TNF-α expression and ROS production. Furthermore, in vitro study proved a similar effect of 2-BP on the regulation of Drp1 and OPA1 expression. 2-BP also increased the mitochondrial membrane potential and decreased the levels of BAX, ROS, and proinflammatory cytokines. These results indicate that 2-BP may attenuate the inflammatory pain of CFA-treated rats via regulating mitochondrial fission/fusion balance and function.     

Guard cell-specific inhibition of Arabidopsis MPK3 expression causes abnormal stomatal responses to abscisic acid and hydrogen peroxide

MAP kinases have been linked to guard cell signalling. Arabidopsis thaliana MAP Kinase 3 (MPK3) is known to be activated by abscisic acid (ABA) and hydrogen peroxide (H(2)O(2)), which also control stomatal movements. We therefore studied the possible role of MPK3 in guard cell signalling through guard cell-specific antisense inhibition of MPK3 expression. Such transgenic plants contained reduced levels of MPK3 mRNA in the guard cells and displayed partial insensitivity to ABA in inhibition of stomatal opening, but responded normally to this hormone in stomatal closure. However, ABA-induced stomatal closure was reduced compared with controls when cytoplasmic alkalinization was prevented with sodium butyrate. MPK3 antisense plants were less sensitive to exogenous H(2)O(2), both in inhibition of stomatal opening and in promotion of stomatal closure, thus MPK3 is required for the signalling of this compound. ABA-induced H(2)O(2) synthesis was normal in these plants, indicating that MPK3 probably acts in signalling downstream of H(2)O(2). These results provide clear evidence for the important role of MPK3 in the perception of ABA and H(2)O(2) in guard cells.     

Nitric oxide production occurs after cytosolic alkalinization during stomatal closure induced by abscisic acid

Abscisic acid (ABA) raised the cytosolic pH and nitric oxide (NO) levels in guard cells while inducing stomatal closure in epidermis of Pisum sativum. Butyrate (a weak acid) reduced the cytosolic pH/NO production and prevented stomatal closure by ABA. Methylamine (a weak base) enhanced the cytosolic alkalinization and aggravated stomatal closure by ABA. The rise in guard cell pH because of ABA became noticeable after 6 min and peaked at 12 min, while NO production started at 9 min and peaked at 18 min. These results suggested that NO production was downstream of the rise in cytosolic pH. The ABA-induced increase in NO of guard cells and stomatal closure was prevented by 2-phenyl-4,4,5,5-tetramethyl imidazoline-1-oxyl 3-oxide (cPTIO, a NO scavenger) and partially by N-nitro-L-Arg-methyl ester (L-NAME, an inhibitor of NO synthase). In contrast, cPTIO or L-NAME had only a marginal effect on the pH rise induced by ABA. Ethylene glycol tetraacetic acid (EGTA, a calcium chelator) prevented ABA-induced stomatal closure while restricting cytosolic pH rise and NO production. We suggest that during ABA-induced stomatal closure, a rise in cytosolic pH is necessary for NO production. Calcium may act upstream of cytosolic alkalinization and NO production, besides its known function as a downstream component.     

Jasmonates Induce Intracellular Alkalinization and Closure ofPaphiopedilumGuard Cells

Jasmonates (jasmonic acid or methyl jasmonate) promote stomatalclosure inPaphiopedilum Supersuk(RHS, 1973) andP. tonsum(Rchb.f)Stein. Studies on guard cells loaded with pH dependent fluorescentdyes show that jasmonates cause intracellular alkalinizationof up to 0.5 pH units within 5 to 15 min. Jasmonate-inducedalkalinization always preceded stomatal closure and where alkalinizationwas not detected no closure occurred. Propionic acid inhibitedjasmonate-induced stomatal closure, suggesting that jasmonate-inducedintracellular alkalinization is involved in guard cell movements. BCECF; confocal microscopy; cytosolic pH; guard cells; jasmonic acid; methyl jasmonate; Paphiopedilum Supersuk(R. H. S.); Snarf-1; stomatal movements     

共聚焦显微技术研究ABA诱导蚕豆气孔保卫细胞H_2O_2的产生(简报)

逆境下,植物细胞内ABA含量急剧增加,同时植物也可通过一些酶代谢反应积累活性氧,如H_2O_2,O_2~-。ABA作为逆境信号对气孔运动的显著调节作用已被诸多实验所证实,但关于其对气孔运动调节的细节还知之甚少。H_2O_2作为氧化信号分子在植物抗病信号转导中已得到广泛研究,但H_2O_2是否介导保卫细胞的气孔运动还缺乏直接的证据。我们已初步发现H_2O_2可参与外源ABA诱     

Calcium-Activated K+ Channels and Calcium-Induced Calcium Release by Slow Vacuolar Ion Channels in Guard Cell Vacuoles Implicated in the Control of Stomatal Closure

Stomatal closing requires the efflux of K+ from the large vacuolar organelle into the cytosol and across the plasma membrane of guard cells. More than 90% of the K+ released from guard cells during stomatal closure originates from the guard cell vacuole. However, the corresponding molecular mechanisms for the release of K+ from guard cell vacuoles have remained unknown. Rises in the cytoplasmic Ca2+ concentration have been shown to trigger ion efflux from guard cells, resulting in stomatal closure. Here, we report a novel type of largely voltage-independent K+-selective ion channel in the vacuolar membrane of guard cells that is activated by physiological increases in the cytoplasmic Ca2+ concentration. These vacuolar K+ (VK) channels had a single channel conductance of 70 pS with 100 mM KCI on both sides of the membrane and were highly selective for K+ over NH4+ and Rb+. Na+, Li+, and Cs+ were not measurably permeant. The Ca2+, voltage, and pH dependences, high selectivity for K+, and high density of VK channels in the vacuolar membrane of guard cells suggest a central role for these K+ channels in the initiation and control of K+ release from the vacuole to the cytoplasm required for stomatal closure. The activation of K+-selective VK channels can shift the vacuolar membrane to more positive potentials on the cytoplasmic side, sufficient to activate previously described slow vacuolar cation channels (SV-type). Analysis of the ionic selectivity of SV channels demonstrated a Ca2+ over K+ selectivity (permeability ratio for Ca2+ to K+ of ~3:1) of these channels in broad bean guard cells and red beet vacuoles, suggesting that SV channels play an important role in Ca2+-induced Ca2+ release from the vacuole during stomatal closure. A model is presented suggesting that the interaction of VK and SV channel activities is crucial in regulating vacuolar K+ and Ca2+ release during stomatal closure. Furthermore, the possibility that the ubiquitous SV channels may represent a general mechanism for Ca2+-induced Ca2+ release from higher plant vacuoles is discussed.     

共聚焦显微技术研究ABA诱导蚕豆气孔保卫细胞H2O2的产生（简报）

The methods of confocal laser scanning microscopy (CLSM) and microinjection were used to study ABA-induced H2O2 in guard cells (Vicia faba), which were labeled with H2O2 specific probe-2, 7-dichlorofluorescin diacetate(H2DCFDA). The results indicated 100 U/mL catalase (CAT) could inhibit partly stomatal closure induced by ABA. 10(-3) mmol/L ABA could significantly induce H2O2 production in chloroplast in guard cells of Vicia faba following microinjection, and 100 U/mL CAT could partly abolish the effects following simultaneous microinjection of ABA and CAT. These suggest that H2O2 is possibly involved in ABA signaling leading to stomatal closure.     

NO可能作为H2O2的下游信号介导ABA诱导的蚕豆气孔关闭

ABA、H2O2和硝普钠(SNP)均能诱导蚕豆气孔关闭.NO的清除剂c-PTIO可以减轻由ABA或H2O2所诱导的蚕豆气孔关闭的程度,而过氧化氢酶(CAT)则不能减轻NO诱导的气孔关闭程度.激光共聚焦显微检测结果显示,10μmo1/L的ABA处理后,胞内H2O2的产生速率明显高于NO的产生速率;CAT几乎可完全抑制ABA所诱导的DAF的荧光增加;外源H2O2能显著诱导胞内DAF的荧光增加;c-PTIO对ABA诱导的DCF荧光略有促进作用,但外源SNP不能诱导胞内DCF荧光增加.这些结果表明,在ABA诱导气孔关闭过程中,H2O2可能在NO的上游起作用并受NO的负反馈调节.     

Cytoplasmic alkalization precedes reactive oxygen species production during methyl jasmonate- and abscisic acid-induced stomatal closure

Signaling events during abscisic acid (ABA) or methyl jasmonate (MJ)-induced stomatal closure were examined in Arabidopsis wild type, ABA-insensitive (ost1-2), and MJ-insensitive mutants (jar1-1) in order to examine a crosstalk between ABA and MJ signal transduction. Some of the experiments were performed on epidermal strips of Pisum sativum. Stomata of jar1-1 mutant plants are insensitive to MJ but are able to close in response to ABA. However, their sensitivity to ABA is less than that of wild-type plants. Reciprocally, the stomata of ost1-2 are insensitive to ABA but are able to close in response to MJ to a lesser extent compared to wild-type plants. Both MJ and ABA promote H(2)O(2) production in wild-type guard cells, while exogenous application of diphenylene iodonium (DPI) chloride, an inhibitor of NAD(P)H oxidases, results in the suppression of ABA- and MJ-induced stomatal closure. ABA elevates H(2)O(2) production in wild-type and jar1-1 guard cells but not in ost1-2, whereas MJ induces H(2)O(2) production in both wild-type and ost1-2 guard cells, but not in jar1-1. MJ-induced stomatal closing is suppressed in the NAD(P)H oxidase double mutant atrbohD/F and in the outward potassium channel mutant gork1. Furthermore, MJ induces alkalization in guard cell cytosol, and MJ-induced stomatal closing is inhibited by butyrate. Analyses of the kinetics of cytosolic pH changes and reactive oxygen species (ROS) production show that the alkalization of cytoplasm precedes ROS production during the stomatal response to both ABA and MJ. Our results further indicate that JAR1, as OST1, functions upstream of ROS produced by NAD(P)H oxidases and that the cytoplasmic alkalization precedes ROS production during MJ or ABA signal transduction in guard cells.     

过氧化氢对蚕豆气孔运动和质膜K+通道的影响

不同浓度H2 O2 可使蚕豆 (ViciafabaL .)叶片气孔关闭 ,抑制气孔张开 ,10mmol/L的H2 O2 最有效 ,10 μmol/L的H2 O2 仍明显使气孔关闭。且 10 μmol/L的H2 O2 抑制气孔张开作用能被EGTA所消除 ,表明Ca2 参与低浓度H2 O2 使气孔关闭的过程。 2mmol/L的H2 O2 可使质膜内向K 通道电流明显减小 ,而外向K 通道电流显著增加。因此 ,H2 O2 促进蚕豆气孔关闭主要是通过抑制K 通过保卫细胞质膜内向流入 ,或加强K 外向流出实现的     

Intra-vacuolar reserves of membranes during stomatal closure: the possible role of guard cell vacuoles estimated by 3-D reconstruction

Stomatal apertures are regulated by morphological changes in guard cells which have been associated with guard cell vacuolar structures. To investigate the contribution of guard cell vacuoles to stomatal movement, we examined the dynamics of vacuolar membrane structures in guard cells and evaluated the changes in vacuolar volumes and surface areas during stomatal movement. Using a transgenic Arabidopsis line expressing green fluorescent protein (GFP)-AtVAM3, we have found that the guard cell vacuolar structures became complicated during stomatal closure with the appearance of numerous intra-vacuolar membrane structures. A three-dimensional (3-D) reconstruction using our originally developed software, REANT (reconstructor and analyzer of 3-D structure), and photobleaching analysis revealed the continuity of the vacuolar structures, even when they appeared to be compartmented in confocal images of closed stomata. Furthermore, calculations of the surface area by REANT revealed an increase in vacuolar surface area during stomatal closure but a decrease in the surface area of the guard cells. Movement of a vital staining dye, FM4-64, to the vacuolar membrane was accelerated during ABA-induced stomatal closure in Vicia faba. These results suggest that the guard cell vacuoles store some portion of the excess membrane materials produced during stomatal closure as intra-vacuolar structures.