ROS-independent Mechanisms Research Articles

Carbonic Anhydrase inhibitors bearing organotelluride moieties as novel agents for antitumor therapy

Solid tumors are mainly characterized by a specific hypoxic microenvironment which makes them particularly challenging to treat. The Carbonic Anhydrase IX (CA IX) is one of the major enzymes implicated in the regulation and maintaining of such conditions and therefore its targeting represents a winning approach in recent tumor targeted therapy. In our search for an innovative combination therapy, we attained the synthesis of selective CA IX inhibitors which are also used for cell specific delivery of cytotoxic organotellurium scaffolds. We investigated compounds 5b, 7b and 7c for their redox properties by means of radical species scavenging and lipid peroxidation inhibitory capacity, as well as intracellular (reactive oxygen species) ROS production in both normal and cancer cell lines. Subsequently, compounds were evaluated as possible free radical generators by ESR spectrometry showing to cause or promote the formation of free radicals. These results accounted for a novel, potent, and selective CA IX inhibitor (i.e. 7c, Ki = 32 nM) with high cytotoxic effect against malignant melanoma (MeWo) and hepatocellular carcinoma (HepG2) cells over normal fibroblasts (NHDF) through ROS-independent mechanisms. The preliminary data gives support to employ organotellurium moieties as useful pharmacological tools for further development in the oncological field.

N-acetylcysteine Can Induce Massive Oxidative Stress, Resulting in Cell Death with Apoptotic Features in Human Leukemia Cells.

N-acetylcysteine (NAC), often used as an antioxidant-scavenging reactive oxygen species (ROS) in vitro, was recently shown to increase the cytotoxicity of other compounds through ROS-dependent and ROS-independent mechanisms. In this study, NAC itself was found to induce extensive ROS production in human leukemia HL-60 and U937 cells. The cytotoxicity depends on ROS-modulating enzyme expression. In HL-60 cells, NAC activated NOX2 to produce superoxide (O2•−). Its subsequent conversion into H2O2 by superoxide dismutase 1 and 3 (SOD1, SOD3) and production of ClO− from H2O2 by myeloperoxidase (MPO) was necessary for cell death induction. While the addition of extracellular SOD potentiated NAC-induced cell death, extracellular catalase (CAT) prevented cell death in HL-60 cells. The MPO inhibitor partially reduced the number of dying HL-60 cells. In U937 cells, the weak cytotoxicity of NAC is probably caused by lower expression of NOX2, SOD1, SOD3, and by the absence of MOP expression. However, even here, the addition of extracellular SOD induced cell death in U937 cells, and this effect could be reversed by extracellular CAT. NAC-induced cell death exhibited predominantly apoptotic features in both cell lines. Conclusions: NAC itself can induce extensive production of O2•− in HL-60 and U937 cell lines. The fate of the cells then depends on the expression of enzymes that control the formation and conversion of ROS: NOX, SOD, and MPO. The mode of cell death in response to NAC treatment bears apoptotic and apoptotic-like features in both cell lines.

New Insights on Hemp Oil Enriched in Cannabidiol: Decarboxylation, Antioxidant Properties and In Vitro Anticancer Effect.

This study aimed to obtain and characterize extracted hemp oil enriched in cannabidiol (CBD) by decarboxylation of cannabidiolic acid (CBDA) and to give new insights into its antioxidant and anticancer effects. Optimization of CBDA decarboxylation in hemp oil was performed, and CBD and CBDA contents and purities were determined by flash chromatography, 1H- and 13C-NMR. The antioxidant properties of CBD-enriched oil were investigated by Fe2+ chelating activity, Fe3+ reducing antioxidant power assay, O2●− scavenging activity, HO● scavenging ability and lipid peroxidation inhibitory assay, and its cytotoxicity, apoptosis- and oxidative stress-inducing effects on NHDF, MeWo, HeLa, HepG2 and HOS cells were determined. The CBD concentration in hemp oil was increased by CBDA soft decarboxylation optimized at 90 °C, for 1 h and the resulting oil was capable of reducing iron, scavenging free radicals and inhibiting lipid peroxidation in cell-free oxidative conditions. CBD-enriched oil promoted NHDF proliferation at up to 15 µg CBD/mL, while inducing apoptosis and ROS production and modulating antioxidant enzymes’ gene expression in cancer cells, being selective for osteosarcoma cells, and induced apoptosis by p53- and ROS-independent mechanisms. CBD-enriched hemp oil demonstrated antioxidant properties in oxidative conditions and promoted normal fibroblasts’ proliferation, while inducing apoptosis and ROS production in cancer cells.

PM2.5 and the typical components cause organelle damage, apoptosis and necrosis: Role of reactive oxygen species

In this research, the organelle damage, apoptosis and necrosis induced by PM2.5, BC and Kaolin were studied using human bronchial epithelial (16HBE) cells. PM2.5, BC and Kaolin all induce cell death, LDH release and excess intracellular ROS generation. For the organelle injuries, Kaolin and high-dose PM2.5 (240 μg/mL) cause lysosomal acidification, but BC causes lysosomal alkalization (lysosomal membrane permeabilization, LMP). BC and Kaolin cause the loss of mitochondrial membrane potential (MMP), while PM2.5 does not. For the cell death mode, PM2.5 causes both apoptosis and necrosis. However only necrosis has been detected in the BC and Kaolin treated groups, indicating the more severe cellular insult. Excess ROS generation is involved in the organelle damage and cell death. ROS contributes to the BC-induced LMP and necrosis, but does not significantly affect the Kaolin-induced MMP loss and necrosis. Therefore, the BC component in PM2.5 may cause cytotoxicity via ROS-dependent pathways, the Kaolin component may damage cells via ROS-independent mechanisms such as strong interaction. The PM2.5-induced apoptosis and necrosis can be partially mitigated after the removal of ROS, indicating the existence of both the ROS-dependent and ROS-independent mechanisms due to the complicated PM2.5 components. BC represents the anthropogenic source component in PM2.5, while Kaolin represents the natural source component. Our results provide knowledge on the toxic mechanisms of typical PM2.5 components at the cellular and subcellular levels.

Noxious effects of selected food-occurring oxidized amino acids on differentiated CACO-2 intestinal human cells

The harmful effects of food-occurring oxidized amino acids, namely, aminoadipic acid (AAA), dityrosine (DTYR), L-kynurenine (KN), kynurenic acid (KA) and 3-nitrotyrosine (3NT), were studied on differentiated CACO-2 cells by flow cytometry and quantification of glutathione (GSH), and allysine. Cells were exposed to food-relevant doses (200 μM) of each compound for 4 or 72h and compared to a control (no stimulated cells). All oxidized amino acids induced apoptosis and results indicated that underlying mechanisms depended on the chemical nature of the species. AAA, KN and KA caused ROS generation and severe oxidative stress in 96%, 98% and 89% of exposed cells (77% in control cells), leading to significant GSH depletion and allysine accretion (1.5, 1.5 and 1.6 nmol allysine/mg protein, respectively at 4h; control: 0.22 nmol/mg protein; p < 0.05). DTYR and 3NT induced significant apoptosis to 29% and 25% of cells (control: 16%; p < 0.05) and necrosis to 28% and 26% of cells (control: 23%) at 72h by ROS-independent mechanisms. KN and KA were found to induce a cycle arrest effect on CACO-2 cells. These findings emphasize the potential harmful effects of the intake of oxidized proteins and amino acids and urge the necessity of carrying out further molecular studies.

Role of DJ-1 in Immune and Inflammatory Diseases.

The DJ-1 protein, known as an oxidative stress sensor, participates in the onset of oxidative stress-related diseases such as cancer, neurodegenerative disorders, type 2 diabetes, and male infertility. Although DJ-1 has been extensively studied for more than two decades, evidence has only recently emerged that it plays a key role in immune and inflammatory disorders. The immune regulatory function of DJ-1 is achieved by modulating the activation of several immune cells including macrophages, mast cells, and T cells via reactive oxygen species (ROS)-dependent and/or ROS-independent mechanisms. This review describes the current knowledge on DJ-1, focusing on its immune and inflammatory regulatory roles, and highlights the significance of DJ-1 as a novel therapeutic target for immune and inflammatory diseases.

Concentration-dependent cytokine responses of silica nanoparticles and role of ROS in human lung epithelial cells.

Reactive oxygen species (ROS) is regarded as a critical denominator in nanoparticle toxicology and inflammation. Previously, we have shown that silica nanoparticles sized 50nm (Si50) induce release of CXCL8 and IL-6 from BEAS-2B cells, via mechanisms involving NFκB, p38 MAP kinase and TGF-α-activated EGF receptor. In the present study, the role of ROS-mediated mechanisms in the concentration-dependent Si50 induction of CXCL8 and IL-6 responses was examined. Si50 (200µg/mL) induced a time-dependent ROS formation and a postponed increase in expression of haem oxygenase (HO-1) mRNA and protein. Pre-treatment with the ROS inhibitors N-acetyl cysteine (NAC) and diphenyleneiodonium (DPI) partially attenuated CXCL8 and IL-6 responses to 200µg/mL, but not to 100µg/mL Si50. The release of TGF-α induced by Si50 (200µg/mL) was significantly reduced by NAC, but not by DPI nor siRNA against NADPH oxidase DUOX-1 (siDUOX-1). Furthermore, siDUOX-1 reduced Si50-induced CXCL8, but not IL-6. Both p38 and p65 phosphorylations were inhibited by siDUOX-1, but for NAC only p65 phosphorylation reached a significant reduction. Neither NAC nor DPI reduced Si50-induced CXCL8 and IL-6 gene expressions. In conclusion, Si50-induced CXCL8 and IL-6 involved both ROS-dependent and ROS-independent mechanisms. Notably, the role of ROS seemed restricted to effects of higher concentrations of Si50 and not mediated via the gene expression.

Antimycin A-induced mitochondrial dysfunction activates vagal sensory neurons via ROS-dependent activation of TRPA1 and ROS-independent activation of TRPV1

Inflammation causes activation of nociceptive sensory nerves, resulting in debilitating sensations and reflexes. Inflammation also induces mitochondrial dysfunction through multiple mechanisms. Sensory nerve terminals are densely packed with mitochondria, suggesting that mitochondrial signaling may play a role in inflammation-induced nociception. We have previously shown that agents that induce mitochondrial dysfunction, such as antimycin A, activate a subset of nociceptive vagal sensory nerves that express transient receptor potential (TRP) channels ankyrin 1 (A1) and vanilloid 1 (V1). However, the mechanisms underlying these responses are incompletely understood. Here, we studied the contribution of TRPA1, TRPV1 and reactive oxygen species (ROS) to antimycin A-induced vagal sensory nerve activation in dissociated neurons and at the sensory terminals of bronchopulmonary C-fibers. Nociceptive neurons were defined chemically and genetically. Antimycin A-evoked activation of vagal nociceptors in a Fura2 Ca2+ assay correlated with TRPV1 responses compared to TRPA1 responses. Nociceptor activation was dependent on both TRP channels, with TRPV1 predominating in a majority of responding nociceptors and TRPA1 predominating only in nociceptors with the greatest responses. Surprisingly, both TRPA1 and TRPV1 were activated by H2O2 when expressed in HEK293. Nevertheless, targeting ROS had no effect of antimycin A-evoked TRPV1 activation in either HEK293 or vagal neurons. In contrast, targeting ROS inhibited antimycin A-evoked TRPA1 activation in HEK293, vagal neurons and bronchopulmonary C-fibers, and a ROS-insensitive TRPA1 mutant was completely insensitive to antimycin A. We therefore conclude that mitochondrial dysfunction activates vagal nociceptors by ROS-dependent (TRPA1) and ROS-independent (TRPV1) mechanisms.

Characterization of metabolic profiles and lipopolysaccharide effects on porcine vascular wall mesenchymal stem cells.

The link between metabolic remodeling and stem cell fate is still unclear. To explore this topic, the metabolic profile of porcine vascular wall mesenchymal stem cells (pVW-MSCs) was investigated. At the first and second cell passages, pVW-MSCs exploit both glycolysis and cellular respiration to synthesize adenosine triphosphate (ATP), but in the subsequent (third to eighth) passages they do not show any mitochondrial ATP turnover. Interestingly, when the first passage pVW-MSCs are exposed to 0.1 or 10 μg/ml lipopolysaccharides (LPSs) for 4 hr, even if ATP synthesis is prevented, the spare respiratory capacity is retained and the glycolytic capacity is unaffected. In contrast, the exposure of pVW-MSCs at the fifth passage to 10 μg/ml LPS stimulates mitochondrial ATP synthesis. Flow cytometry rules out any reactive oxygen species (ROS) involvement in the LPS effects, thus suggesting that the pVW-MSC metabolic pattern is modulated by culture conditions via ROS-independent mechanisms.

The Role of Iron in Friedreich's Ataxia: Insights From Studies in Human Tissues and Cellular and Animal Models.

Friedreich’s ataxia (FRDA) is a rare early-onset degenerative disease that affects both the central and peripheral nervous systems, and other extraneural tissues, mainly the heart and endocrine pancreas. This disorder progresses as a mixed sensory and cerebellar ataxia, primarily disturbing the proprioceptive pathways in the spinal cord, peripheral nerves and nuclei of the cerebellum. FRDA is an inherited disease with an autosomal recessive pattern caused by an insufficient amount of the nuclear-encoded mitochondrial protein frataxin, which is an essential and highly evolutionary conserved protein whose deficit results in iron metabolism dysregulation and mitochondrial dysfunction. The first experimental evidence connecting frataxin with iron homeostasis came from Saccharomyces cerevisiae; iron accumulates in the mitochondria of yeast with deletion of the frataxin ortholog gene. This finding was soon linked to previous observations of iron deposits in the hearts of FRDA patients and was later reported in animal models of the disease. Despite advances made in the understanding of FRDA pathophysiology, the role of iron in this disease has not yet been completely clarified. Some of the questions still unresolved include the molecular mechanisms responsible for the iron accumulation and iron-mediated toxicity. Here, we review the contribution of the cellular and animal models of FRDA and relevance of the studies using FRDA patient samples to gain knowledge about these issues. Mechanisms of mitochondrial iron overload are discussed considering the potential roles of frataxin in the major mitochondrial metabolic pathways that use iron. We also analyzed the effect of iron toxicity on neuronal degeneration in FRDA by reactive oxygen species (ROS)-dependent and ROS-independent mechanisms. Finally, therapeutic strategies based on the control of iron toxicity are considered.

Differential anti-tumour effects of MTH1 inhibitors in patient-derived 3D colorectal cancer cultures

Reactive oxygen species (ROS) function as second messengers in signal transduction, but high ROS levels can also cause cell death. MTH1 dephosphorylates oxidized nucleotides, thereby preventing their incorporation into DNA and protecting tumour cells from oxidative DNA damage. Inhibitors of MTH1 (TH588 and (S)-crizotinib) were shown to reduce cancer cell viability. However, the MTH1-dependency of the anti-cancer effects of these drugs has recently been questioned. Here, we have assessed anti-tumour effects of TH588 and (S)-crizotinib in patient-derived 3D colorectal cancer cultures. Hypoxia and reoxygenation – conditions that increase intracellular ROS levels – increased sensitivity to (S)-crizotinib, but not to TH588. (S)-crizotinib reduced tyrosine phosphorylation of c-MET and ErbB3 whereas TH588 induced a mitotic cell cycle arrest, which was not affected by adding ROS-modulating compounds. Furthermore, we show that both compounds induced DNA damage that could not be prevented by adding the ROS inhibitor N-acetyl-L-cysteine. Moreover, adding ROS-modulating compounds did not alter the reduction in viability in response to TH588 and (S)-crizotinib. We conclude that TH588 and (S)-crizotinib have very clear and distinct anti-tumour effects in 3D colorectal cancer cultures, but that these effects most likely occur through distinct and ROS-independent mechanisms.

Neutrophil extracellular traps prevent HIV infection in the female genital tract.

Women acquire HIV mainly through sexual intercourse. However, low transmission rates per sexual act indicate that local immune mechanisms contribute to HIV prevention. Neutrophils represent 10-20% of the genital immune cells in healthy women. Neutrophils mediate mucosal protection against bacterial and fungal pathogens through different mechanisms, including the release of Neutrophil Extracellular Traps (NETs). NETs are DNA fragments associated with antimicrobial granular proteins. Despite neutrophil abundance and central contributions to innate immunity in the genital tract, their role in protection against HIV-acquisition is unknown. We found that stimulation of human genital neutrophils with HIV viral-like particles (HIV-VLP) induced NET release within minutes of viral exposure, through ROS-independent mechanisms that resulted in immediate entrapment of HIV-VLPs. Incubation of infectious HIV with pre-formed genital NETs prevented infection of susceptible cells through irreversible viral inactivation. HIV inactivation by NETs from genital neutrophils could represent a previously unrecognized form of mucosal protection against HIV-acquisition.

Mitochondrial modulation-induced activation of vagal sensory neuronal subsets by antimycin A, but not CCCP or rotenone, correlates with mitochondrial superoxide production.

Inflammation causes nociceptive sensory neuron activation, evoking debilitating symptoms and reflexes. Inflammatory signaling pathways are capable of modulating mitochondrial function, resulting in reactive oxygen species (ROS) production, mitochondrial depolarization and calcium release. Previously we showed that mitochondrial modulation with antimycin A, a complex III inhibitor, selectively stimulated nociceptive bronchopulmonary C-fibers via the activation of transient receptor potential (TRP) ankyrin 1 (A1) and vanilloid 1 (V1) cation channels. TRPA1 is ROS-sensitive, but there is little evidence that TRPV1 is activated by ROS. Here, we used dual imaging of dissociated vagal neurons to investigate the correlation of mitochondrial superoxide production (mitoSOX) or mitochondrial depolarization (JC-1) with cytosolic calcium (Fura-2AM), following mitochondrial modulation by antimycin A, rotenone (complex I inhibitor) and carbonyl cyanide m-chlorophenyl hydrazone (CCCP, mitochondrial uncoupling agent). Mitochondrial modulation by all agents selectively increased cytosolic calcium in a subset of TRPA1/TRPV1-expressing (A1/V1+) neurons. There was a significant correlation between antimycin A-induced calcium responses and mitochondrial superoxide in wild-type ‘responding’ A1/V1+ neurons, which was eliminated in TRPA1-/- neurons, but not TRPV1-/- neurons. Nevertheless, antimycin A-induced superoxide production did not always increase calcium in A1/V1+ neurons, suggesting a critical role of an unknown factor. CCCP caused both superoxide production and mitochondrial depolarization but neither correlated with calcium fluxes in A1/V1+ neurons. Rotenone-induced calcium responses in ‘responding’ A1/V1+ neurons correlated with mitochondrial depolarization but not superoxide production. Our data are consistent with the hypothesis that mitochondrial dysfunction causes calcium fluxes in a subset of A1/V1+ neurons via ROS-dependent and ROS-independent mechanisms.

Recent advances in the mechanism of selenoamino acids toxicity in eukaryotic cells.

Selenium is an essential trace element due to its incorporation into selenoproteins with important biological functions. However, at high doses it is toxic. Selenium toxicity is generally attributed to the induction of oxidative stress. However, it has become apparent that the mode of action of seleno-compounds varies, depending on its chemical form and speciation. Recent studies in various eukaryotic systems, in particular the model organism Saccharomyces cerevisiae, provide new insights on the cytotoxic mechanisms of selenomethionine and selenocysteine. This review first summarizes current knowledge on reactive oxygen species (ROS)-induced genotoxicity of inorganic selenium species. Then, we discuss recent advances on our understanding of the molecular mechanisms of selenocysteine and selenomethionine cytotoxicity. We present evidences indicating that both oxidative stress and ROS-independent mechanisms contribute to selenoamino acids cytotoxicity. These latter mechanisms include disruption of protein homeostasis by selenocysteine misincorporation in proteins and/or reaction of selenols with protein thiols.

Dynamic Redox Regulation of IL-4 Signaling.

Quantifying the magnitude and dynamics of protein oxidation during cell signaling is technically challenging. Computational modeling provides tractable, quantitative methods to test hypotheses of redox mechanisms that may be simultaneously operative during signal transduction. The interleukin-4 (IL-4) pathway, which has previously been reported to induce reactive oxygen species and oxidation of PTP1B, may be controlled by several other putative mechanisms of redox regulation; widespread proteomic thiol oxidation observed via 2D redox differential gel electrophoresis upon IL-4 treatment suggests more than one redox-sensitive protein implicated in this pathway. Through computational modeling and a model selection strategy that relied on characteristic STAT6 phosphorylation dynamics of IL-4 signaling, we identified reversible protein tyrosine phosphatase (PTP) oxidation as the primary redox regulatory mechanism in the pathway. A systems-level model of IL-4 signaling was developed that integrates synchronous pan-PTP oxidation with ROS-independent mechanisms. The model quantitatively predicts the dynamics of IL-4 signaling over a broad range of new redox conditions, offers novel hypotheses about regulation of JAK/STAT signaling, and provides a framework for interrogating putative mechanisms involving receptor-initiated oxidation.

Abstract 272: Chronic Hyperlipidemia Alters the Population of Endothelial Progenitor Cells in Bone Marrow and Peripheral Circulation via Both ROS-dependent and Independent Mechanisms

Background/Aims: Bone marrow (BM)-derived endothelial progenitor cells (EPCs) make significant contribution to the function and integrity of vasculature. The number of EPCs is significantly decreased in hyperlipidemic patients. Reactive oxygen species (ROS) and oxidative stress were considered an important mechanism for the development of atherosclerosis in hyperlipidemia. The present study was to determine the role of ROS production in the changes of EPC population in chronic hyperlipidemia. Methods and Results: EPC numbers and ROS formation in BM and blood were determined in wild-type (WT) male C57BL/6 mice and hyperlipidemic LDL receptor knockout (LDLR-/-) mice with high fat diet for 4 months. Intracellular blood, extracellular BM and blood ROS production was significantly increased in hyperlipidemic LDLR-/- mice that was effectively blocked with N-acetylcysteine treatment. Hyperlipidemia produced complex changes in EPC populations in BM and blood. The c-Kit+/CD31+ cell number was significantly decreased in BM and blood, and the numbers of CD34+/CD133+ cells and Sca-1+/Flk-1+ cells were significantly decreased in blood without change in BM, which were not affected by inhibition of ROS production. Interestingly, blood CD34+/Flk-1+ cell number was significantly increased in hyperlipidemic mice that was prevented when ROS formation was inhibited. Conclusions: Chronic hyperlipidemia produced significant and complex changes in EPC populations in both BM and circulation through both ROS-dependent and ROS-independent mechanisms in mice.

Mitochondrial and cytoplasmic ROS have opposing effects on lifespan.

Reactive oxygen species (ROS) are highly reactive, oxygen-containing molecules that can cause molecular damage within the cell. While the accumulation of ROS-mediated damage is widely believed to be one of the main causes of aging, ROS also act in signaling pathways. Recent work has demonstrated that increasing levels of superoxide, one form of ROS, through treatment with paraquat, results in increased lifespan. Interestingly, treatment with paraquat robustly increases the already long lifespan of the clk-1 mitochondrial mutant, but not other long-lived mitochondrial mutants such as isp-1 or nuo-6. To genetically dissect the subcellular compartment in which elevated ROS act to increase lifespan, we deleted individual superoxide dismutase (sod) genes in clk-1 mutants, which are sensitized to ROS. We find that only deletion of the primary mitochondrial sod gene, sod-2 results in increased lifespan in clk-1 worms. In contrast, deletion of either of the two cytoplasmic sod genes, sod-1 or sod-5, significantly decreases the lifespan of clk-1 worms. Further, we show that increasing mitochondrial superoxide levels through deletion of sod-2 or treatment with paraquat can still increase lifespan in clk-1;sod-1 double mutants, which live shorter than clk-1 worms. The fact that mitochondrial superoxide can increase lifespan in worms with a detrimental level of cytoplasmic superoxide demonstrates that ROS have a compartment specific effect on lifespan – elevated ROS in the mitochondria acts to increase lifespan, while elevated ROS in the cytoplasm decreases lifespan. This work also suggests that both ROS-dependent and ROS-independent mechanisms contribute to the longevity of clk-1 worms.

Fcγ and Complement Receptors and Complement Proteins in Neutrophil Activation in Rheumatoid Arthritis: Contribution to Pathogenesis and Progression and Modulation by Natural Products.

Rheumatoid arthritis (RA) is a highly disabling disease that affects all structures of the joint and significantly impacts on morbidity and mortality in RA patients. RA is characterized by persistent inflammation of the synovial membrane lining the joint associated with infiltration of immune cells. Eighty to 90% of the leukocytes infiltrating the synovia are neutrophils. The specific role that neutrophils play in the onset of RA is not clear, but recent studies have evidenced that they have an important participation in joint damage and disease progression through the release of proteolytic enzymes, reactive oxygen species (ROS), cytokines, and neutrophil extracellular traps, in particular during frustrated phagocytosis of immune complexes (ICs). In addition, the local and systemic activation of the complement system contributes to the pathogenesis of RA and other IC-mediated diseases. This review discusses (i) the participation of Fcγ and complement receptors in mediating the effector functions of neutrophils in RA; (ii) the contribution of the complement system and ROS-dependent and ROS-independent mechanisms to joint damage in RA; and (iii) the use of plant extracts, dietary compounds, and isolated natural compounds in the treatment of RA, focusing on modulation of the effector functions of neutrophils and the complement system activity and/or activation.

Inhibition of early upstream events in prodromal Alzheimer's disease by use of targeted antioxidants.

A link between Alzheimer's disease (AD) and an excess presence of oxidant free radicals in the brain has frequently been reported. It is generally assumed that such oxidative stress and related cellular damage is caused by inflammatory changes in the brain and is consequent to amyloid deposition. This review makes the argument that elevated oxidative stress in AD is an early causal event in the initiation and advancement of this disease. Oxidative stress can be decreased by enhancing antioxidant enzymes through activation of the cytoplasmic transcriptional factor (Nrf2)/ARE (antioxidant response element) pathway, and by dietary and endogenous antioxidant chemicals. Reduction in the binding ability of Nrf2 to ARE lowers antioxidant enzyme levels. Decreased levels of Nrf2 and augmentation of oxidative stress in AD suggest that the ROS-dependent mechanism of activating the Nrf2/ARE pathway has become unresponsive. A combination of agents that can either activate the Nrf2-ARE pathway by ROS-independent mechanisms, or by acting directly as antioxidant chemicals, may be necessary to reduce oxidative stress in AD. Earlier shortcomings of using individual antioxidants may be due to consideration of antioxidants as pharmacological agents, ignoring the fact that individual antioxidants can be transmuted in the highly oxidant milieu that is present in AD. Interactions between various cellular compartments may require simultaneous examination of more than one agent. The clinical utility of such a more integrative method can reveal interactive effects such as those found in nutritional research and this can compensate for any mechanistic shortcomings of simultaneous testing of more than a single agent.

SAH-induced MMP activation and K V current suppression is mediated via both ROS-dependent and ROS-independent mechanisms.

Voltage-gated potassium (K V) channels regulate cerebral artery tone and have been implicated in subarachnoid hemorrhage (SAH)-induced pathologies. Here, we examined whether matrix metalloprotease (MMP) activation contributes to SAH-induced K V current suppression and cerebral artery constriction via activation of epidermal growth factor receptors (EGFRs). Using patch clamp electrophysiology, we observed that K V currents were selectively decreased in cerebral artery myocytes isolated from SAH model rabbits. Consistent with involvement of enhanced MMP and EGFR activity in SAH-induced K V current suppression, we found that: (1) oxyhemoglobin (OxyHb) and/or the exogenous EGFR ligand, heparin-binding EGF-like growth factor (HB-EGF), failed to induce further K V current suppression after SAH and (2) gelatin zymography detected significantly higher MMP-2 activity after SAH. The removal of reactive oxygen species (ROS) by combined treatment with superoxide dismutase (SOD) and catalase partially inhibited OxyHb-induced K V current suppression. However, these agents had little effect on OxyHb-induced MMP-2 activation. Interestingly, in the presence of a broad-spectrum MMP inhibitor (GM6001), OxyHb failed to cause K V current suppression. These data suggest that OxyHb suppresses K V currents through both ROS-dependent and ROS-independent pathways involving MMP activation. The ROS-independent pathway involves activation of MMP-2, whereas the ROS-dependent pathway involves activation of a second unidentified MMP or ADAM (a disintegrin and metalloprotease domain).