活性氧中间体和NO在植物抗病中的作用

植物与病原菌互作时,活性氧中间体(reactiveoxygenintermediates,ROI)和一氧化氮(NO)参与了植物抗病性的建立。寄主与病原菌非亲合性互作产生二次氧爆发高峰,体内NO增加。许多氧化酶可以催化氧爆发产生ROI。ROI和NO通过氧化还原信号启动寄主细胞局部的过敏性坏死反应和全株系统获得性抗病性。     

活性氧中间体和NO在植物抗病中的作用

植物与病原菌互作时,活性氧中间体（reactive oxygen intermediates,ROI）和一氧化氮(NO)参与了植物抗病性的建立。寄主与病原菌非亲合性互作产生二次氧爆发高峰,体内NO增加。许多氧化酶可以催化氧爆发产生ROI。ROI和NO通过氧化还原信号启动寄主细胞局部的过敏性坏死反应和全株系统获得性抗病性。     

Oleoyl-Lysophosphatidylcholine Limits Endothelial Nitric Oxide Bioavailability by Induction of Reactive Oxygen Species

Previously we reported modulation of endothelial prostacyclin and interleukin-8 production, cyclooxygenase-2 expression and vasorelaxation by oleoyl- lysophosphatidylcholine (LPC 18:1). In the present study, we examined the impact of this LPC on nitric oxide (NO) bioavailability in vascular endothelial EA.hy926 cells. Basal NO formation in these cells was decreased by LPC 18:1. This was accompanied with a partial disruption of the active endothelial nitric oxide synthase (eNOS)- dimer, leading to eNOS uncoupling and increased formation of reactive oxygen species (ROS). The LPC 18:1-induced ROS formation was attenuated by the superoxide scavenger Tiron, as well as by the pharmacological inhibitors of eNOS, NADPH oxidases, flavin-containing enzymes and superoxide dismutase (SOD). Intracellular ROS-formation was most prominent in mitochondria, less pronounced in cytosol and undetectable in endoplasmic reticulum. Importantly, Tiron completely prevented the LPC 18:1-induced decrease in NO bioavailability in EA.hy926 cells. The importance of the discovered findings for more in vivo like situations was analyzed by organ bath experiments in mouse aortic rings. LPC 18:1 attenuated the acetylcholine-induced, endothelium dependent vasorelaxation and massively decreased NO bioavailability. We conclude that LPC 18:1 induces eNOS uncoupling and unspecific superoxide production. This results in NO scavenging by ROS, a limited endothelial NO bioavailability and impaired vascular function.     

一氧化氮对渗透胁迫下小麦种子萌发及其活性氧代谢的影响

一氧化氮供体硝普钠(Sodium nitroprusside,SNP)能明显地促进渗透胁迫下小麦(Triticum aestivum L.)种子萌发、胚根和胚芽伸长,提高萌发过程中淀粉酶和内肽酶的活力,加速贮藏物质的降解：胁迫解除后,仍能使种子维持较高的活力。此外,SNP还能显著诱导渗透胁迫下CAT、APX活力的上升和脯氨酸含量积累,抑制LOX活力,从而提高渗透胁迫下小麦种子萌发过程中抗氧化能力。进一步研究还发现,SNP诱导切胚半粒小麦种子萌发早期(6h)的淀粉酶活力上升可能与GA3无直接关系。     

一氧化氮对渗透胁迫下小麦种子萌发及其活性氧代谢的影响

一氧化氮供体硝普钠(Sodium nitroprusside,SNP)能明显地促进渗透胁迫下小麦(Triticum aestivum L.)种子萌发、胚根和胚芽伸长,提高萌发过程中淀粉酶和内肽酶的活力,加速贮藏物质的降解;胁迫解除后,仍能使种子维持较高的活力.此外,SNP还能显著诱导渗透胁迫下CAT、APX活力的上升和脯氨酸含量积累,抑制LOX活力,从而提高渗透胁迫下小麦种子萌发过程中抗氧化能力.进一步研究还发现,SNP诱导切胚半粒小麦种子萌发早期(6h)的淀粉酶活力上升可能与GA3无直接关系.     

Nitric Oxide Is Required for Homeostasis of Oxygen and Reactive Oxygen Species in Barley Roots under Aerobic Conditions

Oxygen, the terminal electron acceptor for mitochondrial electron transport, is vital for plants because of its role in the production of ATP by oxidative phosphorylation. While photosynthetic oxygen production contributes to the oxygen supply in leaves, reducing the risk of oxygen limitation of mitochondrial metabolism under most conditions, root tissues often suffer oxygen deprivation during normal development due to the lack of an endogenous supply and isolation from atmospheric oxygen.     

Functional Interaction of ROS and Nitric Oxide during Induction of Heat Resistance of Wheat Seedlings by Hydrogen Sulfide Donor

Russian Journal of Plant Physiology - The participation of reactive oxygen species (ROS) and nitric oxide (NO), and also enzymatic systems generating them, in the development of heat resistance of...     

Interplay Among Nitric Oxide and Reactive Oxygen Species: A Complex Network Determining Cell Survival or Death

Programmed cell death (PCD) is an integrated cellular process occurring in plant growth, development, and defense responses to facilitate normal growth and development and better survival against various stresses as a whole. As universal toxic chemicals in plant and animal cells, reactive oxygen or nitrogen species (ROS or RNS), mainly superoxide anion (O₂^−•), hydrogen peroxide (H₂O₂) or nitric oxide (^•NO), have been studied extensively for their roles in PCD induction. Physiological and genetic studies have convincingly shown their essential roles. However, the details and mechanisms by which ROS and ^•NO interplay and induce PCD are not well understood. Our recent study on Cupressus lusitanica culture cell death revealed the elicitor-induced co-accumulation of ROS and ^•NO and interactions between ^•NO and H₂O₂ or O₂-^• in different ways to regulate cell death. ^•NO and H₂O₂ reciprocally enhanced the production of each other whereas ^•NO and O₂^−• showed reciprocal suppression on each other''s production. It was the interaction between ^•NO and O₂-^• but not between ^•NO and H₂O₂ that induced PCD, probably through peroxynitrite (ONOO⁻). In this addendum, some unsolved issues in the study were discussed based on recent studies on the complex network of ROS and ^•NO leading to PCD in animals and plants.Key Words: cell death, nitric oxide, reactive oxygen species, interaction, posttranslational modification     

Reactive Oxygen Species: Participation in Cellular Processes and Progression of Pathology

Russian Journal of Bioorganic Chemistry - Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are not only side products of chemical reactions, but participants in various cellular...     

NMDA Receptor Activation Produces Concurrent Generation of Nitric Oxide and Reactive Oxygen Species: Implications for Cell Death

Abstract: The ability of glutamate to stimulate generation of intracellular oxidant species was determined by microfluorescence in cerebellar granule cells loaded with the oxidant-sensitive fluorescent dye 2,7-dichlorofluorescin (DCF). Exposure of cells to glutamate (10 µM) produced a rapid generation of oxidants that was blocked ～70% by MK-801 (a noncompetitive NMDA-receptor antagonist). To determine if nitric oxide (NO) or reactive oxygen species (ROS) contributed to the oxidation of DCF, cells were treated with compounds that altered their generation. NO production was inhibited with N^G-nitro-l -arginine methyl ester (l -NAME) (nitric oxide synthase inhibitor) and reduced hemoglobin (NO scavenger). Alternatively, cells were incubated with superoxide dismutase (SOD) and catalase, which selectively metabolize O₂^?· andH₂O₂. Concurrent inhibition of O₂^?· and NO production nearly abolished intracellular oxidant generation. Pretreatment of cells with either chelerythrine (1 µM, protein kinase C inhibitor) or quinacrine (5 µM, phospholipase A₂ inhibitor) before addition of glutamate also blocked oxidation of DCF. Generation of oxidants by glutamate was significantly reduced by incubating the cells in Ca²⁺-free buffer. In cytotoxicity studies, a positive correlation was observed between glutamate-induced death and oxidant generation. Glutamate-induced cytotoxicity was blocked by MK-801 and attenuated by treatment with l -NAME, chelerythrine, SOD, or quinacrine. It is concluded that glutamate induces concurrent generation of NO and ROS by activation of both NMDA receptors and non-NMDA receptors through a Ca²⁺-mediated process. Activation of NO synthase and phospholipaseA₂ contribute significantly to this response. It is proposed that simultaneous generation of NO and ROS results in formation of peroxynitrite, which initiates the cellular damage.     

活性氧参与-氧化氮诱导的神经细胞凋亡

采用激光共聚焦成像技术,用氧化还原敏感的特异性荧光探针(DCFH-DA和DHR123)直接研究了一氧 化氮供体S-亚硝基-N-乙酰基青霉胺(SNAP)诱导未成熟大鼠小脑颗粒神经元凋亡过程中的细胞胞浆、线粒体 中活性氧水平的变化,发现神经细胞经0.5mmol/LSNAP处理1h后,细胞胞浆及线粒体中活性氧水平大大增 加.一氧化氮清除剂血红蛋白能够有效抑制细胞胞浆、线粒体中活性氧的产生,防止细胞凋亡.外源性谷胱甘 肽对细胞也具有良好的保护作用,而当细胞中谷胱甘肽的合成被抑制后,一氧化氮的神经毒性大大增强.实验 结果表明一氧化氮通过促进神经细胞产生内源性活性氧而启动细胞凋亡程序,而谷胱甘肽可能是重要的防止一 氧化氮引发神经损伤的内源性抗氧化剂     

Insulin Reverses D-Glucose–Increased Nitric Oxide and Reactive Oxygen Species Generation in Human Umbilical Vein Endothelial Cells

Vascular tone is controlled by the L-arginine/nitric oxide (NO) pathway, and NO bioavailability is strongly affected by hyperglycaemia-induced oxidative stress. Insulin leads to high expression and activity of human cationic amino acid transporter 1 (hCAT-1), NO synthesis and vasodilation; thus, a protective role of insulin on high D-glucose–alterations in endothelial function is likely. Vascular reactivity to U46619 (thromboxane A₂ mimetic) and calcitonin gene related peptide (CGRP) was measured in KCl preconstricted human umbilical vein rings (wire myography) incubated in normal (5 mmol/L) or high (25 mmol/L) D-glucose. hCAT-1, endothelial NO synthase (eNOS), 42 and 44 kDa mitogen-activated protein kinases (p42/44^mapk), protein kinase B/Akt (Akt) expression and activity were determined by western blotting and qRT-PCR, tetrahydrobiopterin (BH₄) level was determined by HPLC, and L-arginine transport (0–1000 μmol/L) was measured in response to 5–25 mmol/L D-glucose (0–36 hours) in passage 2 human umbilical vein endothelial cells (HUVECs). Assays were in the absence or presence of insulin and/or apocynin (nicotinamide adenine dinucleotide phosphate-oxidase [NADPH oxidase] inhibitor), tempol or Mn(III)TMPyP (SOD mimetics). High D-glucose increased hCAT-1 expression and activity, which was biphasic (peaks: 6 and 24 hours of incubation). High D-glucose–increased maximal transport velocity was blocked by insulin and correlated with lower hCAT-1 expression and SLC7A1 gene promoter activity. High D-glucose–increased transport parallels higher reactive oxygen species (ROS) and superoxide anion (O₂ ^•–) generation, and increased U46619-contraction and reduced CGRP-dilation of vein rings. Insulin and apocynin attenuate ROS and O₂ ^•– generation, and restored vascular reactivity to U46619 and CGRP. Insulin, but not apocynin or tempol reversed high D-glucose–increased NO synthesis; however, tempol and Mn(III)TMPyP reversed the high D-glucose–reduced BH₄ level. Insulin and tempol blocked the high D-glucose–increased p42/44^mapk phosphorylation. Vascular dysfunction caused by high D-glucose is likely attenuated by insulin through the L-arginine/NO and O₂ ^•–/NADPH oxidase pathways. These findings are of interest for better understanding vascular dysfunction in states of foetal insulin resistance and hyperglycaemia.     

Stimulation of p42 and p44 Mitogen-Activated Protein Kinases by Reactive Oxygen Species and Nitric Oxide in Hippocampus

Abstract: Reactive oxygen species (ROS) have been suggested to act as cellular messengers that mediate signal transduction cascades in various cell types. However, little is known about their role in this capacity in the nervous system. We have begun to investigate the role of ROS, and that of nitric oxide (NO), in mediating mitogen-activated protein kinase (MAPK) signaling in rat hippocampal slices. Our studies have revealed that direct exposure of hippocampal slices to hydrogen peroxide, xanthine/xanthine oxidase (a superoxide-generating system), sodium nitroprusside (an NO donor compound), S -nitroso- N -acetylpenicillamine (an NO donor compound), or 3-morpholinosydnonimine (a compound that produces NO and superoxide) results in an enhancement in tyrosine phosphorylation of several proteins, including proteins with apparent molecular masses of 42 and 44 kDa. We investigated the possibility that these proteins correspond to the active forms of p42 MAPK and p44 MAPK. Hippocampal slices exposed to various ROS and NO donors resulted in increases in levels of the active forms of both p42 MAPK and p44 MAPK. The ROS- and NO-enhanced tyrosine phosphorylation and activation of p42 MAPK and p44 MAPK were inhibited by pretreatment with the antioxidant N -acetyl- l -cysteine. Our observations indicate that ROS and NO can mediate protein tyrosine phosphorylation and MAPK signaling in the hippocampus via a redox-sensitive mechanism and suggest a potential cellular mechanism for their effects in the nervous system.     

活性氧参与一氧化氮诱导的神经细胞凋亡

采用激光共聚焦成像技术，用氧化还原敏感的特异性荧光探针(DCFH-DA和DHR123)直接研究了一氧化氮供体S-亚硝基-N-乙酰基青霉胺(SNAP)诱导未成熟大鼠小脑颗粒神经元凋亡过程中的细胞胞浆、线粒体中活性氧水平的变化，发现神经细胞经0.5 mmol/L SNAP处理1 h后，细胞胞浆及线粒体中活性氧水平大大增加.一氧化氮清除剂血红蛋白能够有效抑制细胞胞浆、线粒体中活性氧的产生，防止细胞凋亡.外源性谷胱甘肽对细胞也具有良好的保护作用，而当细胞中谷胱甘肽的合成被抑制后，一氧化氮的神经毒性大大增强.实验结果表明一氧化氮通过促进神经细胞产生内源性活性氧而启动细胞凋亡程序，而谷胱甘肽可能是重要的防止一氧化氮引发神经损伤的内源性抗氧化剂.     

The Role of Nitric Oxide and Reactive Oxygen Species in the Killing of Leishmania braziliensis by Monocytes from Patients with Cutaneous Leishmaniasis

Human cutaneous leishmaniasis (CL) caused by Leishmania braziliensis, presents an exaggerated Th1 response that is associated with ulcer development. Macrophages are the primary cells infected by Leishmania parasites and both reactive oxygen species (ROS) and nitric oxide (NO) are important in the control of Leishmania by these cells. The mechanism involved in the killing of L. braziliensis is not well established. In this study, we evaluate the role of ROS and NO in the control of L. braziliensis infection by monocytes from CL patients. After in vitro infection with L. braziliensis, the oxidative burst by monocytes from CL patients was higher when compared to monocytes from healthy subjects (HS). Inhibition of the ROS pathway caused a significant decrease in the oxidative burst in L. braziliensis infected monocytes from both groups. In addition, we evaluated the intracellular expression of ROS and NO in L. braziliensis-infected monocytes. Monocytes from CL patients presented high expression of ROS after infection with L. braziliensis. The expression of NO was higher in monocytes from CL patients as compared to expression in monocytes from HS. A strong positive correlation between NO production and lesion size of CL patients was observed. The inhibition of ROS production in leishmania-infected monocytes from CL patients allowed the growth of viable promastigotes in culture supernatants. Thus, we demonstrate that while production of ROS is involved in L. braziliensis killing, NO alone is not sufficient to control infection and may contribute to the tissue damage observed in human CL.     

Rehydration of the Lichen Ramalina lacera Results in Production of Reactive Oxygen Species and Nitric Oxide and a Decrease in Antioxidants

Lichens are slow-growing associations of fungi and unicellular green algae or cyanobacteria. They are poikilohydric organisms whose lifestyle in many cases consists of alternating periods of desiccation, with low metabolic activity, and hydration, which induces increase in their metabolism. Lichens have apparently adapted to such extreme transitions between desiccation and rehydration, but the mechanisms that govern these adaptations are still poorly understood. In this study, the effect of rehydration on the production of reactive oxygen species and nitric oxide as well as low-molecular-weight antioxidants was investigated with the lichen Ramalina lacera. Rehydration of R. lacera resulted in the initiation of and a rapid increase in photosynthetic activity. Recovery of photosynthesis was accompanied by bursts of intracellular production of reactive oxygen species and nitric oxide. Laser-scanning confocal microscopy using dichlorofluorescein fluorescence revealed that formation of reactive oxygen species following rehydration was associated with both symbiotic partners of the lichen. The rate and extent of reactive oxygen species production were similar in the light and in the dark, suggesting a minor contribution of photosynthesis. Diaminofluorescein fluorescence, indicating nitric oxide formation, was detected only in fungal hyphae. Activities associated with rehydration did not have a deleterious effect on membrane integrity as assessed by measurement of electrolyte leakage, but water-soluble low-molecular-weight antioxidants decreased significantly.