Reactive oxygen species, mitochondria, apoptosis and aging

In this paper, we shall review various antioxygen defense systems of the cell paying particular attention to those that prevent superoxide formation rather than scavenge already formed superoxide and its products. The role of uncoupled, decoupled and non-coupled respiration, mitochondrial pore, mitochondrion-linked apoptosis will be considered. Mitochondrial theory of aging will be regarded in context of reactive oxygen species-induced damage of mitochondrial DNA. (Mol Cell Biochem 174: 305–319, 1997)     

Reactive oxygen species generation and antioxidant systems in plant mitochondria

In living cells, reactive oxygen species (ROS) play a key role in signaling but these compounds can also damage macromolecules. As in other compartments, the mitochondrial ROS concentrations need to be tightly controlled. Plant mitochondria contain several antioxidant systems that are not only able to scavenge ROS and limit their production but also to repair damages to macromolecules and possibly to serve as redox sensors. They comprise ascorbate- and glutathione-dependent pathways as well as systems based on thioredoxin (TRX)- and glutaredoxin (GRX)-like molecules. This review describes the various mitochondrial redox pathways for ROS control in plants with special emphasis on the poorly studied GRX and TRX systems and provides perspectives for future research in this area.     

Reactive oxygen species in regulation of fungal development

Reactive oxygen species (ROS) are formed by fungi in the course of metabolic activity. ROS production increases in fungi due to various stress agents such as starvation, light, mechanical damage, and interactions with some other living organisms. Regulation of ROS level appears to be very important during development of the fungal organism. ROS sources in fungal cells, their sensors, and ROS signal transduction pathways are discussed in this review. Antioxidant defense systems in different classes of fungi are characterized in detail. Particular emphasis is placed on ROS functions in interactions of phytopathogenic fungi with plant cells.     

Reactive oxygen species regulate DNA copy number in isolated yeast mitochondria by triggering recombination-mediated replication

Mitochondrial DNA (mtDNA) encodes proteins that are essential for cellular ATP production. Reactive oxygen species (ROS) are respiratory byproducts that damage mtDNA and other cellular components. In Saccharomyces cerevisiae, the oxidized base excision-repair enzyme Ntg1 introduces a double-stranded break (DSB) at the mtDNA replication origin ori5; this DSB initiates the rolling-circle mtDNA replication mediated by the homologous DNA pairing protein Mhr1. Thus, ROS may play a role in the regulation of mtDNA copy number. Here, we show that the treatment of isolated mitochondria with low concentrations of hydrogen peroxide increased mtDNA copy number in an Ntg1- and Mhr1-dependent manner. This treatment elevated the DSB levels at ori5 of hypersuppressive [rho^–] mtDNA only if Ntg1 was active. In vitro Ntg1-treatment of hypersuppressive [rho^–] mtDNA extracted from hydrogen peroxide-treated mitochondria revealed increased oxidative modifications at ori5 loci. We also observed that purified Ntg1 created breaks in single-stranded DNA harboring oxidized bases, and that ori5 loci have single-stranded character. Furthermore, chronic low levels of hydrogen peroxide increased in vivo mtDNA copy number. We therefore propose that ROS act as a regulator of mtDNA copy number, acting through the Mhr1-dependent initiation of rolling-circle replication promoted by Ntg1-induced DSB in the single-stranded regions at ori5.     

Production of reactive oxygen species by mitochondria: central role of complex III

The mitochondrial respiratory chain is a major source of reactive oxygen species (ROS) under pathological conditions including myocardial ischemia and reperfusion. Limitation of electron transport by the inhibitor rotenone immediately before ischemia decreases the production of ROS in cardiac myocytes and reduces damage to mitochondria. We asked if ROS generation by intact mitochondria during the oxidation of complex I substrates (glutamate, pyruvate/malate) occurred from complex I or III. ROS production by mitochondria of Sprague-Dawley rat hearts and corresponding submitochondrial particles was studied. ROS were measured as H2O2 using the amplex red assay. In mitochondria oxidizing complex I substrates, rotenone inhibition did not increase H2O2. Oxidation of complex I or II substrates in the presence of antimycin A markedly increased H2O2. Rotenone prevented antimycin A-induced H2O2 production in mitochondria with complex I substrates but not with complex II substrates. Catalase scavenged H2O2. In contrast to intact mitochondria, blockade of complex I with rotenone markedly increased H2O2 production from submitochondrial particles oxidizing the complex I substrate NADH. ROS are produced from complex I by the NADH dehydrogenase located in the matrix side of the inner membrane and are dissipated in mitochondria by matrix antioxidant defense. However, in submitochondrial particles devoid of antioxidant defense ROS from complex I are available for detection. In mitochondria, complex III is the principal site for ROS generation during the oxidation of complex I substrates, and rotenone protects by limiting electron flow into complex III.     

细胞内活性氧与肿瘤

活性氧是细胞癌变过程中的重要角色：它本身能使DNA损伤,同时又促使致癌物质的产生并且许多致癌因子都是先诱导产生活性氧,然后通过活性氧起致癌作用的,但是另一方面,活性氧却有杀伤癌细胞和诱导细胞凋亡的能力许多抗癌药正是利用活性氧这一特点起作用的。     

Reactive oxygen species and the regulation of hyperproliferation in a colonial hydroid

Colonies of Podocoryna carnea circulate gastrovascular fluid among polyps via tubelike stolons. At polyp-stolon junctions, mitochondrion-rich cells in part regulate this gastrovascular flow. During competition, colonies hyperproliferate nematocytes and stolons; nematocysts are discharged until one colony is killed. Hyperproliferation then ceases, and normal growth resumes. Here, competing colonies were treated with azide, which inhibits respiration and upregulates reactive oxygen species (ROS). After the cessation of competition, azide-treated colonies continued to hyperproliferate. In azide-treated competing colonies, however, mitochondrion-rich cells were found to produce similar amounts of ROS as those in untreated competing colonies. Subsequent experiments showed that both azide treatment and competition diminished the lumen widths at polyp-stolon junctions, where mitochondrion-rich cells are found. In competing colonies, these diminished widths may also diminish the metabolic demand on these cells, causing mitochondria to enter the resting state and emit more ROS. Indeed, results with two fluorescent probes show that mitochondrion-rich cells in competing colonies produce more ROS than those in noncompeting colonies. In sum, these results suggest that competition perturbs the usual activity of mitochondrion-rich cells, altering their redox state and increasing ROS formation. Via uncharacterized pathways, these ROS may contribute to hyperproliferation.     

Reactive oxygen species from mitochondria mediate SW480 cells apoptosis induced by Na2SeO3

A number of selenium compounds have been found to inhibit tumorigenesis in a variety of animal and cell models. In order to explore the molecular mechanism involved in the anticarcinogenesis activity of selenium, we examined the effects of sodium selenite on cell viabilty, generation of reactive oxygen species (ROS), and mitochondrial transmembrane potential (Δω _m ) in human colonic carcinoma cells SW480. The result from MTT test showed that sodium selenite reduced cell viability. Morophologic and flow cytometric results indicated that Na₂SeO₃ induced the apoptosis of SW480 cells. Na₂SeO₃ increased the generation of intracellular ROS, whereas BAPTA-AM, rotenone, and NaCN completely inhibited the increase of ROS induced by Na₂SeO₃. Na₂SeO₃ also caused the disruption of Δω _m . The intracellular ROS increase and apoptosis induced by Na₂SeO₃ were significantly decreased by superoxide dismutase (SOD), catalase. These data suggest that the ROS mediate apoptosis induced by Na₂SeO₃ and mitochondria may be a major source of Na₂SeO₃-induced ROS.     

Reactive oxygen species are involved in regulation of pollen wall cytomechanics

Production and scavenging of reactive oxygen species (ROS) in somatic plant cells is developmentally regulated and plays an important role in the modification of cell wall mechanical properties. Here we show that H₂O₂ and the hydroxyl radical (^?OH) can regulate germination of tobacco pollen by modifying the mechanical properties of the pollen intine (inner layer of the pollen wall). Pollen germination was affected by addition of exogenous H₂O₂, ^?OH, and by antioxidants scavenging endogenous ROS: superoxide dismutase, superoxide dismutase/catalase mimic Mn‐5,10,15,20‐tetrakis(1‐methyl‐4‐pyridyl)21H, 23H‐porphin, or a spin‐trap α‐(4‐pyridyl‐1‐oxide)‐N‐tert‐butylnitrone, which eliminates ^?OH. The inhibiting concentrations of exogenous H₂O₂ and ^?OH did not decrease pollen viability, but influenced the mechanical properties of the wall. The latter were estimated by studying the resistance of pollen to hypo‐osmotic shock. ^?OH caused excess loosening of the intine all over the surface of the pollen grain, disrupting polar growth induction. In contrast, H₂O₂, as well as partial removal of endogenous ^?OH, over‐tightened the wall, impeding pollen tube emergence. Feruloyl esterase (FAE) was used as a tool to examine whether H₂O₂‐inducible inter‐polymer cross‐linking is involved in the intine tightening. FAE treatment caused loosening of the intine and stimulated pollen germination and pollen tube growth, revealing ferulate cross‐links in the intine. Taken together, the data suggest that pollen intine properties can be regulated differentially by ROS. ^?OH is involved in local loosening of the intine in the germination pore region, while H₂O₂ is necessary for intine strengthening in the rest of the wall through oxidative coupling of feruloyl polysaccharides.     

Reactive oxygen species from smooth muscle mitochondria initiate cold-induced constriction of cutaneous arteries

Cold constricts cutaneous blood vessels by selectively increasing the activity of smooth muscle alpha2-adrenoceptors (alpha2-ARs). In mouse tail arteries, alpha2-AR constriction is mediated by alpha2A-ARs at 37 degrees C, whereas the cold-induced augmentation in alpha2-AR activity is mediated entirely by alpha2C-ARs. Cold causes translocation of alpha2C-ARs from the trans-Golgi to the plasma membrane, mediated by cold-induced activation of RhoA and Rho kinase. The present experiments analyzed the mechanisms underlying these responses. Mouse tail arteries were studied in a pressure myograph. Cooling the arteries (28 degrees C) caused a rapid increase in reactive oxygen species (ROS) in smooth muscle cells, determined by confocal microscopy of arteries loaded with the ROS-sensitive probes, dichlorodihydrofluorescein or reduced Mitotracker Red. The inhibitor of mitochondrial complex I rotenone (10 micromol/l), the antioxidant N-acetylcysteine (NAC; 20 mmol/l), or the cell-permeable mimic of superoxide dismutase MnTMPyP (50 micromol/l) did not affect vasoconstriction to alpha2-AR stimulation (UK-14304) at 37 degrees C but dramatically inhibited the response at 28 degrees C. Indeed, these ROS inhibitors abolished the cold-induced increase in alpha2-AR constrictor activity. NAC (20 mmol/l) or MnTMPyP (50 micromol/l) also abolished the cold-induced activation of RhoA in human cultured vascular smooth muscle cells and the cold-induced mobilization of alpha2C-ARs to the cell surface in human embryonic kidney 293 cells transfected with the receptor. The combined results suggest that cold-induced constriction is mediated by redox signaling in smooth muscle cells, initiated by mitochondrial generation of ROS, which stimulate RhoA/Rho kinase signaling and the subsequent mobilization of alpha2C-ARs to the cell surface. Altered activity of ROS may contribute to cold-induced vasospasm occurring in Raynaud's phenomenon.     

Calcium-induced generation of reactive oxygen species in brain mitochondria is mediated by permeability transition

Mitochondrial uptake of calcium in excitotoxicity is associated with subsequent increase in reactive oxygen species (ROS) generation and delayed cellular calcium deregulation in ischemic and neurodegenerative insults. The mechanisms linking mitochondrial calcium uptake and ROS production remain unknown but activation of the mitochondrial permeability transition (mPT) may be one such mechanism. In the present study, calcium increased ROS generation in isolated rodent brain and human liver mitochondria undergoing mPT despite an associated loss of membrane potential, NADH and respiration. Unspecific permeabilization of the inner mitochondrial membrane by alamethicin likewise increased ROS independently of calcium, and the ROS increase was further potentiated if NAD(H) was added to the system. Importantly, calcium per se did not induce a ROS increase unless mPT was triggered. Twenty-one cyclosporin A analogs were evaluated for inhibition of calcium-induced ROS and their efficacy clearly paralleled their potency of inhibiting mPT-mediated mitochondrial swelling. We conclude that while intact respiring mitochondria possess powerful antioxidant capability, mPT induces a dysregulated oxidative state with loss of GSH- and NADPH-dependent ROS detoxification. We propose that mPT is a significant cause of pathological ROS generation in excitotoxic cell death.     

Reactive oxygen and DNA damage in mitochondria.

During the last decade the importance of reactive oxygen species as major contributors to various types of cancer, heart diseases, cataracts, Parkinson's and other degenerative diseases that come with age, and to natural aging has become apparent. Mitochondria are the most important intracellular source of reactive oxygen. Mitochondrial DNA is heavily damaged by reactive oxygen at the bases, as indicated by the high steady-state level of 8-hydroxydeoxyguanosine, the presence of which causes mispairing and point mutations. Mitochondrial DNA is also oxidatively fragmented to a certain extent. Conceivably, such fragmentation relates to deletions found in mitochondrial DNA. Point mutations and deletions have recently been shown to be etiologically linked to several human diseases and natural aging. Future studies should address the causal relationship between mitochondrial dysfunction, production of reactive oxygen species, and aging.     

Reactive oxygen species and yeast apoptosis

Apoptosis is associated in many cases with the generation of reactive oxygen species (ROS) in cells across a wide range of organisms including lower eukaryotes such as the yeast Saccharomyces cerevisiae. Currently there are many unresolved questions concerning the relationship between apoptosis and the generation of ROS. These include which ROS are involved in apoptosis, what mechanisms and targets are important and whether apoptosis is triggered by ROS damage or ROS are generated as a consequence or part of the cellular disruption that occurs during cell death. Here we review the nature of the ROS involved, the damage they cause to cells, summarise the responses of S. cerevisiae to ROS and discuss those aspects in which ROS affect cell integrity that may be relevant to the apoptotic process.     

Reactive oxygen species and oxidative stress

Oxidative stress is defined as an imbalance between the production of reactive oxygen species (ROS) and the antioxidant capacity of the cell. For long, ROS have been considered as harmful by-products of the normal aerobic metabolism process of the mitochondria, implicated in a large variety of diseases. But there are now growing evidences that controlled ROS production also play physiological roles especially in regulating cell redox homeostasis and cell signaling. Biological ROS effects are now well documented. Data show that living organisms have not only adapted themselves to coexist with free radicals but have also developed mechanisms to use them advantageously. However their main sources and mechanisms of action remain poorly described. This review focuses on the main properties of ROS and their paradoxical effects.     

Reactive oxygen species and mitochondrial diseases

A variety of diseases have been associated with excessive reactive oxygen species (ROS), which are produced mostly in the mitochondria as byproducts of normal cell respiration. The interrelationship between ROS and mitochondria suggests shared pathogenic mechanisms in mitochondrial and ROS-related diseases. Defects in oxidative phosphorylation can increase ROS production, whereas ROS-mediated damage to biomolecules can have direct effects on the components of the electron transport system. Here, we review the molecular mechanisms of ROS production and damage, as well as the existing evidence of mitochondrial ROS involvement in human diseases.     

Reactive oxygen species and airway inflammation

Reactive oxygen species may be generated by several inflammatory cells which participate in airway inflammation and their production may be increased in asthma. Oxygen metabolites may contribute to the epithelial damage which is characteristic of asthmatic airways and may activate cells such as mast cells in the airway mucosa. Reactive oxygen species may cause bronchoconstriction, mucus secretion, have effects on airway vasculature, and may increase airway responsiveness. The role of reactive oxygen species in airway disease has been largely neglected, but appears to be an important area for future study. It is also possible that antioxidant defenses may be defective in asthma. If reactive oxygen species participate in the inflammatory response in airway disease, then radical scavengers or antioxidants could play a useful role in therapy.     

Reactive oxygen species and plant immunity

Reactive oxygen species (ROS) play a key role in plant defense mechanisms. They exert direct antimicrobial action, catalyze the mechanical strengthening of cell walls, function as secondary messengers in the superoxide synthase signal pathway and in triggering the hypersensitive response. Although recent studies have unraveled a nature and the mechanisms of the oxidative burst, many questions related to its mode of regulation, its modulation of signaling networks that control growth, development and defense responses remain unanswered.     

Reactive oxygen species and seed germination

Reactive oxygen species (ROS) are continuously produced by the metabolically active cells of seeds, and apparently play important roles in biological processes such as germination and dormancy. Germination and ROS accumulation appear to be linked, and seed germination success may be closely associated with internal ROS contents and the activities of ROS-scavenging systems. Although ROS were long considered hazardous molecules, their functions as cell signaling compounds are now well established and widely studied in plants. In seeds, ROS have important roles in endosperm weakening, the mobilization of seed reserves, protection against pathogens, and programmed cell death. ROS may also function as messengers or transmitters of environmental cues during seed germination. Little is currently known, however, about ROS biochemistry or their functions or the signaling pathways during these processes, which are to be considered in the present review.