Mechanisms Of Reactive Oxygen Species Production Research Articles

Phosphonates-enhanced production of reactive oxygen species by ferrous ion in the presence of oxygen: Effects and mechanisms

Due to their potential environmental hazards, the ongoing release of phosphonates into natural water bodies has sparked serious worries. Phosphonates are typically found as metal-phosphonate complexes in natural water and their complex with Fe2+ is predominant. In this study, we demonstrated that phosphonates could enhance reactive oxygen species (ROS) production upon complexation with Fe2+ in the presence of O2 (from the air). Using aminotris (methyphosphonic acid) (ATMP) as a model phosphonate, we thoroughly investigated the effects and mechanism of ROS production in the ATMP/Fe2+/O2 system. OH•-mediated oxidation was responsible for the transformation of co-existing contaminants. With oxygen (O-) and nitrogen (N-) in ATMP complexed with Fe2+, the redox potential of Fe2+/Fe3+ was significantly reduced, thus promoting OH• production. Further experiments identified OH• production followed the one-electron transfer pathway (O2 → O2•- → H2O2 → OH•). Due to the generation of OH•, 61.5% of phenol (10 mg/L) was degraded and a small fraction of ATMP underwent C-P bond cleavage. The phenomenon of ATMP-enhanced ROS production and co-existing pollutants degradation occurred throughout a wide pH spectrum from 3.0 to 10.0. This study not only clarified how ROS were produced in the ATMP/Fe2+/O2 system, but also provided new ideas for understanding the environmental behavior of phosphonates in promoting co-existing pollutants degradation.

Electrocatalytic Treatment of Pharmaceutical Wastewater by Transition Metals Encapsulated by B, N-Doped CNTs

The electrochemical advanced oxidation process is a promising technology for tackling wastewater pollution, but it suffers from poor pH adaptability and slow catalytic kinetics in a neutral and alkaline environment in a homogeneous system, as well as fast release of metal ions in a heterogeneous system. Herein, a boron- and nitrogen-codoped carbon nanotube-encapsulated transition metal (M@BN-C, M–Co, Cu) cathode with a similar structure was synthesized to explore activity trends and mechanisms. Characteristics of Co@BN-C and Cu@BN-C cathodes were examined and compared with the previously synthesized Fe@BN-C bifunctional cathode. The activity of sulfamethazine (SMT) degradation by the Co@BN-C cathode was higher than both Fe@BN-C and Cu@BN-C at pH = 3 and pH = 7, respectively. However, the activity of Co@BN-C was also higher than that of Cu@BN-C and lower than that of Fe@BN-C at pH = 9. It was observed that •OH and 1O2 were the main reactive oxygen species (ROS) using Co@BN-C and Cu@BN-C cathodes. The Co@BN-C generated the highest •OH for efficient SMT degradation through abundant H2O2 generation, exhibiting the highest catalytic activity compared with the Cu@BN-C cathode. Overall, SMT degradation on the Co@BN-C cathode demonstrated better catalytic performance in real wastewater. This study provided insights into the fundamental catalytic trends and mechanisms of ROS production via the M@BN-C cathode, thus contributing to the development of the M@BN-C cathode for catalytic organic pollutant degradation.

Research progress on reactive oxygen species production mechanisms in tumor sonodynamic therapy

In recent years, because of the growing desire to improve the noninvasiveness and safety of tumor treatments, sonodynamic therapy has gradually become a popular research topic. However, due to the complexity of the therapeutic process, the relevant mechanisms have not yet been fully elucidated. One of the widely accepted possibilities involves the effect of reactive oxygen species. In this review, the mechanism of reactive oxygen species production by sonodynamic therapy (SDT) and ways to enhance the sonodynamic production of reactive oxygen species are reviewed. Then, the clinical application and limitations of SDT are discussed. In conclusion, current research on sonodynamic therapy should focus on the development of sonosensitizers that efficiently produce active oxygen, exhibit biological safety, and promote the clinical transformation of sonodynamic therapy.

Research progress on reactive oxygen species production mechanisms in tumor sonodynamic therapy.

In recent years, because of the growing desire to improve the noninvasiveness and safety of tumor treatments, sonodynamic therapy has gradually become a popular research topic. However, due to the complexity of the therapeutic process, the relevant mechanisms have not yet been fully elucidated. One of the widely accepted possibilities involves the effect of reactive oxygen species. In this review, the mechanism of reactive oxygen species production by sonodynamic therapy (SDT) and ways to enhance the sonodynamic production of reactive oxygen species are reviewed. Then, the clinical application and limitations of SDT are discussed. In conclusion, current research on sonodynamic therapy should focus on the development of sonosensitizers that efficiently produce active oxygen, exhibit biological safety, and promote the clinical transformation of sonodynamic therapy.

Selective induction of rapid cytotoxic effect in glioblastoma cells by oscillating magnetic fields.

The mechanisms underlying anticancer effects of electromagnetic fields are poorly understood. An alternating electric field-generating therapeutic device called Optune™ device has been approved for the treatment of glioblastoma (GBM). We have developed a new device that generates oscillating magnetic fields (OMF) by rapid rotation of strong permanent magnets in specially designed patterns of frequency and timing and have used it to treat an end-stage recurrent GBM patient under an expanded access/compassionate use treatment protocol. Here, we ask whether OMF causes selective cytotoxic effects in GBM and whether it is through generation of reactive oxygen species (ROS). We stimulated patient derived GBM cells, lung cancer cells, normal human cortical neurons, astrocytes, and bronchial epithelial cells using OMF generators (oncoscillators) of our Oncomagnetic Device and compared the results to those obtained under unstimulated or sham-stimulated control conditions. Quantitative fluorescence microscopy was used to assess cell morphology, viability, and ROS production mechanisms. We find that OMF induces highly selective cell death of patient derived GBM cells associated with activation of caspase 3, while leaving normal tissue cells undamaged. The cytotoxic effect of OMF is also seen in pulmonary cancer cells. The underlying mechanism is a marked increase in ROS in the mitochondria, possibly in part through perturbation of the electron flow in the respiratory chain. Rotating magnetic fields produced by a new noninvasive device selectively kill cultured human glioblastoma and non-small cell lung cancer cells by raising intracellular reactive oxygen species, but not normal human tissue cells.

Origin of sonocatalytic activity of fluorescent carbon dots

Although ultrasound-induced reactive oxygen species (ROS) have believed to be primary intermediates for the realization of sonodynamic therapy and sonocatalytic reactions, the mechanism of ROS production under ultrasound has not been well clarified till now. Here we discovered that ultrasound cavitation can break up oxygen-containing groups on the surface of carbon dots (CDs) to generate ROS, giving rise to a novel mechanism different from as-reported those. The bond cleavage of oxygen-containing groups contribute transient oxygen free radicals (•O−) that involve in the subsequent formation of ROS by quickly reacting with hydrogen ions and water, thereby leading to a twice higher sonocatalytic activity of CDs than TiO2. Moreover, the sonosensitizer function of CDs can be tuned by pH values of the surrounding environment that determine the generation and evolution reaction of oxygen free radicals under ultrasound irradiation. This finding paves a new way to design multifunctional sonosensitizers and utilize them in various fields.

Interactions of copper and copper chelate compounds with the amyloid beta peptide: An investigation into electrochemistry, reactive oxygen species and peptide aggregation

Alzheimer's disease is a fatal neurological disorder affecting millions of people worldwide with an increasing patient population as average life expectancy increases. Accumulation of amyloid beta (Aβ) plaques is characteristic of the disease and has been the target of numerous failed clinical trials. In light of this, therapeutics that target mechanisms of neuronal death beyond Aβ aggregation are needed. One potential target is the formation of reactive oxygen species (ROS) that are created during an interaction between Aβ and copper ions. This work shows that ROS production can be slowed by disrupting the interaction between Aβ and copper using copper chelating compounds. We demonstrated that ROS are produced in the presence of Aβ and copper in solution by monitoring H2O2 production using a fluorescence-based assay, which increased when Cu2+ interacted with Aβ. In addition, we were able to show reduced ROS production, without exacerbating the aggregation of Aβ and in some cases alleviating it, by adding copper chelating ligands to the solution. Using cyclic voltammetry, we investigated how these different ligands influenced the electrochemical behavior of copper in solution revealing important insights into the mechanisms of ROS production and chemical interactions that result in decreased ROS rates.

Effect of Reactive Oxygen Species on the Endoplasmic Reticulum and Mitochondria during Intracellular Pathogen Infection of Mammalian Cells.

Oxidative stress, particularly reactive oxygen species (ROS), are important for innate immunity against pathogens. ROS directly attack pathogens, regulate and amplify immune signals, induce autophagy and activate inflammation. In addition, production of ROS by pathogens affects the endoplasmic reticulum (ER) and mitochondria, leading to cell death. However, it is unclear how ROS regulate host defense mechanisms. This review outlines the role of ROS during intracellular pathogen infection, mechanisms of ROS production and regulation of host defense mechanisms by ROS. Finally, the interaction between microbial pathogen-induced ROS and the ER and mitochondria is described.

Mechanisms of Mitochondrial ROS Production in Assisted Reproduction: The Known, the Unknown, and the Intriguing.

The consensus that assisted reproduction technologies (ART), like in vitro fertilization, to induce oxidative stress (i.e., the known) belies how oocyte/zygote mitochondria—a major presumptive oxidative stressor—produce reactive oxygen species (ROS) with ART being unknown. Unravelling how oocyte/zygote mitochondria produce ROS is important for disambiguating the molecular basis of ART-induced oxidative stress and, therefore, to rationally target it (e.g., using site-specific mitochondria-targeted antioxidants). I review the known mechanisms of ROS production in somatic mitochondria to critique how oocyte/zygote mitochondria may produce ROS (i.e., the unknown). Several plausible site- and mode-defined mitochondrial ROS production mechanisms in ART are proposed. For example, complex I catalyzed reverse electron transfer-mediated ROS production is conceivable when oocytes are initially extracted due to at least a 10% increase in molecular dioxygen exposure (i.e., the intriguing). To address the term oxidative stress being used without recourse to the underlying chemistry, I use the species-specific spectrum of biologically feasible reactions to define plausible oxidative stress mechanisms in ART. Intriguingly, mitochondrial ROS-derived redox signals could regulate embryonic development (i.e., their production could be beneficial). Their potential beneficial role raises the clinical challenge of attenuating oxidative damage while simultaneously preserving redox signaling. This discourse sets the stage to unravel how mitochondria produce ROS in ART, and their biological roles from oxidative damage to redox signaling.

Production of Reactive Oxygen Species by the Reaction of Periodate and Hydroxylamine for Rapid Removal of Organic Pollutants and Waterborne Bacteria.

Periodate (PI, IO4-) can be activated by hydroxylamine (HA), resulting in the rapid removal of organic pollutants within seconds. While the previous studies on PI-based advanced oxidation processes (AOPs) have proposed iodate radical (•IO3) as the major reactive species, no evidence of •IO3 production was found in the present PI/HA system. Reactive oxygen species (ROS) including •OH, HO2•, and 1O2 are proposed to be the main oxidants of the PI/HA system, which is supported by various tests employing the scavengers, chemical probes, and spin-trapping electron paramagnetic resonance (EPR) technique. To minimize the risk of toxic iodinated byproduct formation caused by reactive iodine species such as HOI and I2, the molar ratio of HA/PI was optimized at 0.6 to achieve the stoichiometric conversion of IO4- to iodate (IO3-), a preferred nontoxic sink of iodine species. The PI/HA system also efficiently inactivated both Gram-positive and -negative bacteria with producing 1O2 as the dominant disinfectant. The mechanism of ROS production was also investigated and is discussed in detail. This work offers a simple and highly efficient option for PI activation and ROS production which might find useful applications where urgent and rapid removal of toxic pollutants is needed.

Neurotoxicity of Mn3O4 nanoparticles: Apoptosis and dopaminergic neurons damage pathway.

Mn3O4 nanoparticles (NPs) are used increasingly in various fields due to their excellent physiochemical properties. Previous studies have documented that Mn-based nanomaterials resulted in excess reactive oxygen species (ROS) generation and dopamine (DA) reduction both in vivo and in vitro experiments. However, little is known about the mechanism of ROS production and DA decrease induced by Mn-based nanomaterials. The present study was carried out to elucidate the mechanism of the co-incubation model of dopaminergic neuron PC12 cells and the synthesized Mn3O4 NPs. The results demonstrated that exposure to Mn3O4 NPs reduced cell viability, increased level of lactate dehydrogenase (LDH), triggered oxidative stress and induced apoptosis. Notably, the level of ROS was remarkably increased (>10-fold) with Mn3O4 NPs exposure. We also found that mitochondrial calcium Ca2+ uniporter (MCU) was up-regulated and the mitochondrial Ca2+ concentration ([Ca2+]mito) increased induced by Mn3O4 NPs in PC12 cells. Furthermore, the MCU inhibitor RuR significantly attenuated Mn3O4 NPs-induced [Ca2+]mito, ROS production and apoptosis. In PC12 cells, the decrease of DA content was mainly due to the downregulation of DOPA decarboxylase (DDC) expression caused by Mn3O4 NPs treatment. The expression of proteins related to DA storage system was not significantly affected by treatment.

Insufficient fumarase contributes to hypertension by an imbalance of redox metabolism in Dahl salt-sensitive rats.

Fumarase insufficiencies can increase reactive oxygen species (ROS). This study will further dissect the imbalance of redox metabolism and the mechanism of ROS production using proteomic technology in fumarase knockdown HK-2 cells. The contribution of fumarase was further confirmed by supplementation of fumarate and malate in Dahl salt-sensitive rats. Proteomic analysis indicated that fumarase knockdown in HK-2 cells changed the expression or activity of NADPH oxidase (NOX), mitochondrial respiratory chain Complex I and III, ATP synthase subunits, and α-oxoglutarate dehydrogenase (OGDH). Meanwhile, the activities of key antioxidant enzymes, including glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glutathione reductase, glutathione peroxidase, and glutathione S-transferase, increased significantly. The apparent activation of antioxidant defense appeared insufficient as the glutathione and GSH/GSSG ratio were decreased significantly. Dahl salt-sensitive rats exhibited changes in redox metabolism similar to HK-2 cells with fumarase knockdown. Supplementation with fumarate and malate increased and decreased, respectively, blood pressure and H2O2 and malondialdehyde in salt-sensitive rats. These results indicated that insufficient fumarase activity increased ROS by regulating NOX, Complex I and III, ATPase alpha, and OGDH and the imbalance of glutathione metabolism, which may be one of the main reasons for salt-sensitive hypertension. Malate may be a potentially effective drug for the prevention and treatment of salt-sensitive hypertension.

Lysosomotropic challenge of mast cells causes intra-granular reactive oxygen species production

Mast cells contribute to the pathology of allergic and other disorders. Strategies to interfere with harmful mast cell-related activities are therefore warranted. Previously we established a principle for inducing selective apoptosis of mast cells, by the use of lysosomotropic agents that cause secretory granule permeabilization, leading to production of reactive oxygen species (ROS). However, the mechanism of ROS production has not been known. Here we addressed this issue. Live microscopy analysis showed that the secretory granules comprise major subcellular compartments for ROS production in response to mefloquine. As further signs for the primary involvement of secretory granules, both ROS production and cell death was blunted in mast cells lacking serglycin, a secretory granule-restricted proteoglycan. Inhibition of granule acidification caused an essentially complete blockade of granule permeabilization, ROS production and cell death in response to mefloquine. ROS production was also attenuated in the presence of an iron chelator, and after inhibition of either granzyme B or the ERK1/2 MAP kinase signaling pathway. Together, our findings reveal that the mast cell secretory granules constitute major sites for ROS production in mast cells subjected to lysosomotropic challenge. Moreover, this study reveals a central role for granule acidification in ROS generation and the pro-apoptotic response triggered downstream of secretory granule permeabilization.

Oxidative stress in granulosa cells contributes to poor oocyte quality and IVF-ET outcomes in women with polycystic ovary syndrome.

The increased levels of intracellular reactive oxygen species (ROS) in granulosa cells (GCs) may affect the pregnancy results in women with polycystic ovary syndrome (PCOS). In this study, we compared the in vitro fertilization and embryo transfer (IVF-ET) results of 22 patients with PCOS and 25 patients with tubal factor infertility and detected the ROS levels in the GCs of these two groups. Results showed that the PCOS group had significantly larger follicles on the administration day for human chorionic gonadotropin than the tubal factor group (P < 0.05); however, the number of retrieved oocytes was not significantly different between the two groups (P > 0.05). PCOS group had slightly lower fertilization, cleavage, grade I/II embryo, clinical pregnancy, and implantation rates and higher miscarriage rate than the tubal factor group (P > 0.05). We further found a significantly higher ROS level of GCs in the PCOS group than in the tubal factor group (P < 0.05). The increased ROS levels in GCs caused GC apoptosis, whereas NADPH oxidase 2 (NOX2) specific inhibitors (diphenyleneiodonium and apocynin) significantly reduced the ROS production in the PCOS group. In conclusion, the increased ROS expression levels in PCOS GCs greatly induced cell apoptosis, which further affected the oocyte quality and reduced the positive IVF-ET pregnancy results of women with PCOS. NADPH oxidase pathway may be involved in the mechanism of ROS production in GCs of women with PCOS.

Abstract 11614: The Clinical Significance and Novel Mechanism of Reactive Oxygen Species in Heart Failure With Preserved Left Ventricular Ejection Fraction

Introduction: By performing basic and clinical researches, we investigated the clinical significance of reactive oxygen species (ROS) biomarker and the mechanism of ROS production in heart failure with preserved left ventricular ejection fraction (HFpEF). Methods and Results: (1) We measured derivatives of reactive oxidative metabolites (DROM), new biomarker of ROS, in 287 HFpEF patients, and followed them. DROM was significantly higher in HFpEF patients compared to risk factor matched non-HF patients (p&lt;0.001), and was associated with the severity of HF (p&lt;0.001). Kaplan-Meier analysis demonstrated higher probability of cardiovascular events in high-DROM than low-DROM group (p=0.01). Multivariate Cox hazard analysis identified ln-DROM as independent predictor for cardiovascular events (p=0.04). (2) We examined the association of ROS with galectin-3, a predictor of cardiovascular events in HFpEF, by using angiotensin II (AII) receptor blocker (ARB) and galectin-3-inhibitor (Gal3-i) administrated Dahl salt-sensitive rats (DS rats), useful model of HFpEF. Increased left ventricular (LV) galectin-3 protein expressions by Western blotting in HFpEF DS rats were inhibited by ARB, indicating AII-induced galectin-3 expression in HFpEF. Furthermore, Gal3-i significantly reversed LV hypertrophy in DS rats suffered HFpEF (p=0.02), accompanied by the tendency of reduction in collagen-related mRNA gene expression (collagen-I, -III and MMP-2) by real-time RT-PCR. Serum DROM and LV mRNA gene expressions of subunits of ROS generative NADPH oxidase (p22 phox and NOX-2) were increased in HFpEF rats, and Gal3-i tended to downregulate them. These indicated AII induced galectin-3 is involved in LV fibrosis via ROS. (3) Galectin-3 at coronary sinus was higher than at aortic root in HFpEF patients, but not different in non-HF, indicating galectin-3 production in coronary circulation in HFpEF. Moreover, Kaplan-Meier analysis demonstrated higher probability of cardiovascular events in plasma high galectin-3 than in low galectin-3 group in HFpEF patients (p&lt;0.05), indicating clinical significance of galectin-3. Conclusions: These studies demonstrated clinical significance and novel mechanism of ROS in HFpEF.

Abstract A11: Downregulation of NRH:quinone oxidoreductase 2(NQO2) in the LNCaP prostate cancer progression model system: Implications to metastasis

Abstract Prostate cancer is characterized by an innate capacity to produce higher concentrations of reactive oxygen species (ROS). ROS is also a hallmark for highly aggressive disease. Our laboratory has focused on the molecular mechanisms of inhibition of prostate cancer bone metastasis by the naturally occurring plant based anti-oxidant compound curcumin, arising from the rhizomes of Curcuma Longa. These studies highlight the importance of studying the mechanisms of redox homeostasis in normal cells and how they are subverted in cancer cells. The LNCaP prostate cancer progression model system as exemplified by the development of C4, C4-2 and C4-2B ( bone metastatic) sublines derived from the parental LNCaP prostate cancer cells offer an unique avenue to study the role of redox systems in the progression to metastatic disease, culminating in the establishment of osseous metastases. The mechanisms of ROS production and destruction, the cellular machinery and the enzyme systems involved, the relationship of ROS to cellular oncogene activation and the antagonistic duality of oncogenes and tumor suppressor genes with respect to ROS homeostasis have been subjects of intense investigation. How the prostate cancer cells metabolize ROS to achieve their proliferative and metastatic potential is not completely understood. We chose to study one redox system orchestrated by the expression of NADPH:Quinone Oxidoreductase1 (NQO1) and the Nicotinamide Riboside (NRH): Quinone Oxidoreductase 2 (NQO2) in the LNCaP progression model system. Particularly, the enzyme NQO2 has been shown earlier to be the target of another polyphenolic chemopreventive antioxidant resveratrol. The role of NQO2 as a phase II detoxification enzyme acting on several xenobiotics is well characterized. However, the role of NQO2 in regulating ROS and hence cancer progression is only beginning to be understood. With this background, we investigated the potential role of NQO2/NQO1 expression in the transition from androgen dependence to androgen independence and to acquiring bone metastatic potential in C4-2B prostate cancer cells. Our studies indicate that 1) there is a down regulation of NQO2 as the androgen dependent LNCaP cells progress towards androgen independence in C4 and C4-2 cells. In C4-2B cells, there is a barely detectable level of NQO2. In contrast, there is very little change in the level of expression of NQO1; 2) Characteristically, the intracellular ROS production progressively increases from LNCaP to C4-2B; 3) In parallel, there is an up regulation of caveolin-1 as the LNCaP cells progress to bone metastatic C4-2B and 4) a small fraction of NQO2 could be detected in the detergent resistant membrane (DRM) fraction consisting of lipid rafts and caveolae. Our studies suggest that caveolin-1 may play a role in the membrane sequestration of the largely cytoplasmic NQO2 in LNCaP cells and that this interaction could be lost in bone metastatic C4-2B cells. The significance of these findings in relation to the metastatic potential of C4-2B cells will be discussed. Citation Format: Ankeeta Shah, Tze-Chen Hsieh, Rajamma Mathew, Joseph M. Wu, Thambi Dorai. Downregulation of NRH:quinone oxidoreductase 2(NQO2) in the LNCaP prostate cancer progression model system: Implications to metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Metastasis; 2015 Nov 30-Dec 3; Austin, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(7 Suppl):Abstract nr A11.

Differential Roles of the NADPH-Oxidase 1 and 2 in Platelet Activation and Thrombosis.

Reactive oxygen species (ROS) are known to regulate platelet activation; however, the mechanisms of ROS production during platelet activation remain unclear. Platelets express different isoforms of nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) oxidases (NOXs). Here, we investigated the role of NOX1 and NOX2 in ROS generation and platelet activation using NOX1 and NOX2 knockout mice. NOX1(-/Y) platelets showed selective defects in G-protein-coupled receptor-mediated platelet activation induced by thrombin and thromboxane A2 analog U46619, but were not affected in platelet activation induced by collagen-related peptide, a glycoprotein VI agonist. In contrast, NOX2(-/-) platelets showed potent inhibition of collagen-related peptide-induced platelet activation, and also showed partial inhibition of thrombin-induced platelet activation. Consistently, production of ROS was inhibited in NOX1(-/Y) platelets stimulated with thrombin, but not collagen-related peptide, whereas NOX2(-/-) platelets showed reduced ROS generation induced by collagen-related peptide or thrombin. Reduced ROS generation in NOX1/2-deficient platelets is associated with impaired activation of Syk and phospholipase Cγ2, but minimally affected mitogen-activated protein kinase pathways. Interestingly, laser-induced arterial thrombosis was impaired but the bleeding time was not affected in NOX2(-/-) mice. Wild-type thrombocytopenic mice injected with NOX2(-/-) platelets also showed defective arterial thrombosis, suggesting an important role for platelet NOX2 in thrombosis in vivo but not hemostasis. NOX1 and NOX2 play differential roles in different platelet activation pathways and in thrombosis. ROS generated by these enzymes promotes platelet activation via the Syk/phospholipase Cγ2/calcium signaling pathway.

Dynamic changes in the subcellular distribution of the tobacco ROS-producing enzyme RBOHD in response to the oomycete elicitor cryptogein.

Plant NADPH oxidases, also known as respiratory burst oxidase homologues (RBOHs), have been identified as a major source of reactive oxygen species (ROS) during plant-microbe interactions. The subcellular localization of the tobacco (Nicotiana tabacum) ROS-producing enzyme RBOHD was examined in Bright Yellow-2 cells before and after elicitation with the oomycete protein cryptogein using electron and confocal microscopy. The plasma membrane (PM) localization of RBOHD was confirmed and immuno-electron microscopy on purified PM vesicles revealed its distribution in clusters. The presence of the protein fused to GFP was also seen in intracellular compartments, mainly Golgi cisternae. Cryptogein induced, within 1h, a 1.5-fold increase in RBOHD abundance at the PM and a concomitant decrease in the internal compartments. Use of cycloheximide revealed that most of the proteins targeted to the PM upon elicitation were not newly synthesized but may originate from the Golgi pool. ROS accumulation preceded RBOHD transcript- and protein-upregulation, indicating that ROS resulted from the activation of a PM-resident pool of enzymes, and that enzymes newly addressed to the PM were inactive. Taken together, the results indicate that control of RBOH abundance and subcellular localization may play a fundamental role in the mechanism of ROS production.

The Catalytically Active Copper‐Amyloid‐Beta State: Coordination Site Responsible for Reactive Oxygen Species Production

Copper-amyloid-β ROS production: Copper ions (red sphere, see picture) have been found to accumulate in amyloid-β plaques and play a role in the generation of reactive oxygen species (ROS) within this context. Mass spectrometry studies were able to detail the sites of oxidation damage and shed new light on the mechanism of ROS production, important for the understanding of the pathogenicity of amyloid-β peptides.

HCV and oxidative stress in the liver.

Hepatitis C virus (HCV) is the etiological agent accounting for chronic liver disease in approximately 2–3% of the population worldwide. HCV infection often leads to liver fibrosis and cirrhosis, various metabolic alterations including steatosis, insulin and interferon resistance or iron overload, and development of hepatocellular carcinoma or non-Hodgkin lymphoma. Multiple molecular mechanisms that trigger the emergence and development of each of these pathogenic processes have been identified so far. One of these involves marked induction of a reactive oxygen species (ROS) in infected cells leading to oxidative stress. To date, markers of oxidative stress were observed both in chronic hepatitis C patients and in various in vitro systems, including replicons or stable cell lines expressing viral proteins. The search for ROS sources in HCV-infected cells revealed several mechanisms of ROS production and thus a number of cellular proteins have become targets for future studies. Furthermore, during last several years it has been shown that HCV modifies antioxidant defense mechanisms. The aim of this review is to summarize the present state of art in the field and to try to predict directions for future studies.