Oxidative Stress Paradigm Research Articles

Recovery of redox homeostasis altered by CuNPs in H4IIE liver cells does not reduce the cytotoxic effects of these NPs: An investigation using aryl hydrocarbon receptor (AhR) dependent antioxidant activity

The generation of reactive oxygen species (ROS) and consequent oxidative stress is regarded as a relevant mechanism for nanoparticle toxicity. In cells, the activation of the aryl hydrocarbon receptor (AhR) triggers a cascade of defence responses against oxidative stress. By increasing AhR dependent cellular anti-oxidant activity, we tested the extent to which the cytotoxic effect of copper nanoparticles (CuNPs) is governed by oxidative stress. H4IIE rat hepatoma cells were challenged with high ROS levels after exposure to CuNPs, while the AhR-induced cellular anti-oxidant defence was simultaneously activated by the AhR ligand beta-Naphthoflavone (ßNF). Activation of phase II detoxification enzymes (as glutathione-S-transferases, GSTs) and anti-oxidants (glutathione, GSH) led to a complete abrogation of CuNP-induced ROS production. However, a concurrent reduction in cytotoxicity was not detected, thereby indicating that CuNPs exert non-oxidative stress mediated cytotoxic effects. Transmission electron microscopy analysis pointed to a direct physical perturbation of cellular structures by CuNPs, thus contributing to their cytotoxicity. Our observations highlight that distinct mechanisms underlie the toxicity of ions and NPs and indicate that while ROS elicitation is CuNP-specific, the cytotoxic action of these particles may not be directly related to their pro-oxidative activity. These findings have important implications with respect to the oxidative stress paradigm used to explain NP toxicity.

Silica nanoparticle induces oxidative stress and provokes inflammation in human lung cells

Amorphous silica has been used in a wide variety of industrial and consumer applications. Nanosized silica is being considered a potentially ideal nanomaterial for biomedical and biotechnological applications. In the present study, hierarchical oxidative stress paradigms of nanoparticle toxicity were assumed to evaluate the toxicity of amorphous silica nanoparticle in human healthy lung cells, L-132. The cells were exposed to varied concentrations viz. 0, 100, 200, 300, 400 and 500 μg/ml of silica nanoparticle for 24 h. Different parameters of cytotoxicity, oxidative stress and pro-inflammatory markers were assessed. Silica nanoparticle exposure showed concentration-dependent decrease in cell viability, increase in reactive oxygen species with depletion of antioxidant enzyme activity, induction of inteleukin-8 and cyclooxygenase-2 expression and subsequent DNA damage in L-132 cells. The results indicated that amorphous silica nanoparticle followed hierarchical oxidative stress model and generated oxidative stress which provoked the pro-inflammatory response and caused necrosis in human lung cells.

Oxidative stress induced by inorganic nanoparticles in bacteria and aquatic microalgae – state of the art and knowledge gaps

Nanotechnology has revolutionised many areas of modern life, technology and research, which is reflected in the steadily increasing global demand for and consumption of engineered nanomaterials and the inevitable increase of their release into the environment by human activity. The overall long-term impact of engineered nanomaterials on ecosystems is still unknown. Various inorganic nanoparticles have been found to exhibit bactericidal properties and cause growth inhibition in model aquatic microalgae, but the mechanisms of toxicity are not yet fully understood. The causal link between particle properties and biological effects or reactive oxygen species generation is not well established and represents the most eminent quest of nanoecotoxicological investigation. In this review, the current mechanistic understanding of the toxicity of inorganic metal and metal oxide engineered nanomaterials towards bacterial and aquatic microalgal model organisms based on the paradigm of oxidative stress is presented along with a detailed compilation of available literature on the major toxicity factors and research methods.

Mitochondria and Cancer

The future of mitochondrial DNA research has the potential to uncover new insights on genetic diseases and open new opportunities to discover ways to control mitochondria and their influence on the human health and cancer. The outcomes of this work will expand the understanding of cellular respiration and disease risk. In this special issue, we give examples of strategies for the measurement of oxidative stress; a critical factor in tumor progression. While the mitochondrial genome has been well characterized, the associations of the broad spectrum of mitochondrial genotypes remains a relatively rich field of study. Genetic tools are beginning to be realized which characterize mitochondrial populations and link their associations with normal and malignant cells. There are many large-scale deletions which require further investigation. There are populations of genotypes that rise and fall with tissue field effects. Mutations in the mitochondrial genome can alter the cellular biochemical behavior, changing the conditions for potential tumor growth. In this special issue we also address roles of mitochondrial interactions which are central to the physiological processes involved in malignant transformation. Measurements of DNA damage associated with prostate and other cancers can be normalized by comparative measurements of mitochondrial subpopulations. This can be used to assess DNA damage and somatic mutations under physiological and pathological conditions and can serve as a strategy to measure cell toxicity as a guide for devising innovative cancer preventions and treatments. On account that mitochondrial DNA is accessible across various tissues, noninvasive collection and analyses are possible. From such investigations, it has been demonstrated that there is a progression of change in mitochondria through the tissues, as a field effect, that is, associated with the tumor tissue progression. Tumorigenic effects related to increasing ROS are well known in prostate and other solid tumor cancers. Mapping mutations across the entire mitochondrial genome are fundamental to the future work on mitochondrial “omic” investigations. Mitochondrial whole genome sequencing pioneered the concepts of conducting whole genome analyses to understand forensics. This has resulted in efforts to go beyond simple STR typing and to type the entire chromosome. It is because of the increased resolution achieved by sequencing whole metagenomes that, by extension, other fields of diagnostics, personalized medicine, and bacterial and viral forensics have emerged (J. P. Jakupciak unpublished data). Through the study of mitochondria, mechanisms of cancer are emerging. The influence of mitochondria on the metastatic potential of cancer cell lines points to a promising future and in vivo characterization of populations of mitochondria will function as a “looking-glass” into monitoring the modulation events and even predicting changes in metastatic capabilities. Mitochondrial genomics is poised to enhance the over all field of omics and contribute significantly to the advent of personalized medicine [1]. In this special issue, the authors present some of the latest findings in this exciting and rapidly expanding area of genomic research: specific heteroplasmic somatic alterations in the mitochondrial genome contribute to the cell proliferation; a new paradigm for oxidative stress and cell and DNA damage has important implications for both cancer prevention and treatment; an assay for gauging systemic oxidative stress using peripheral blood; upregulation of a nuclear gene whose molecular interactions contribute to mitochondrial dysfunction, promoting cell proliferation; a cancerization field effect described by progressive mitochondrial mutations in noninvasiveand invasive breast cancer. Ryan Parr Andrew Harbottle John P. Jakupciak Gurmit Singh

Use of Metal Oxide Nanoparticle Band Gap To Develop a Predictive Paradigm for Oxidative Stress and Acute Pulmonary Inflammation

We demonstrate for 24 metal oxide (MOx) nanoparticles that it is possible to use conduction band energy levels to delineate their toxicological potential at cellular and whole animal levels. Among the materials, the overlap of conduction band energy (E(c)) levels with the cellular redox potential (-4.12 to -4.84 eV) was strongly correlated to the ability of Co(3)O(4), Cr(2)O(3), Ni(2)O(3), Mn(2)O(3), and CoO nanoparticles to induce oxygen radicals, oxidative stress, and inflammation. This outcome is premised on permissible electron transfers from the biological redox couples that maintain the cellular redox equilibrium to the conduction band of the semiconductor particles. Both single-parameter cytotoxic as well as multi-parameter oxidative stress assays in cells showed excellent correlation to the generation of acute neutrophilic inflammation and cytokine responses in the lungs of C57 BL/6 mice. Co(3)O(4), Ni(2)O(3), Mn(2)O(3), and CoO nanoparticles could also oxidize cytochrome c as a representative redox couple involved in redox homeostasis. While CuO and ZnO generated oxidative stress and acute pulmonary inflammation that is not predicted by E(c) levels, the adverse biological effects of these materials could be explained by their solubility, as demonstrated by ICP-MS analysis. These results demonstrate that it is possible to predict the toxicity of a large series of MOx nanoparticles in the lung premised on semiconductor properties and an integrated in vitro/in vivo hazard ranking model premised on oxidative stress. This establishes a robust platform for modeling of MOx structure-activity relationships based on band gap energy levels and particle dissolution. This predictive toxicological paradigm is also of considerable importance for regulatory decision-making about this important class of engineered nanomaterials.

Effects of Selenium on Calcium Signaling and Apoptosis in Rat Dorsal Root Ganglion Neurons Induced by Oxidative Stress

Ca(2+) is well known for its role as crucial second messenger in modulating many cellular physiological functions, Ca(2+) overload is detrimental to cellular function and may present as an important cause of cellular oxidative stress generation and apoptosis. The aim of this study is to investigate the effects of selenium on lipid peroxidation, reduced glutathione (GSH), glutathione peroxidase (GSH-Px), cytosolic Ca(2+) release, cell viability (MTT) and apoptosis values in dorsal root ganglion (DRG) sensory neurons of rats. DRG cells were divided into four groups namely control, H(2)O(2) (as a model substance used as a paradigm for oxidative stress), selenium, selenium + H(2)O(2). Moderate doses and times of H(2)O(2) and selenium were determined by MTT test. Cells were preterated 200 nM selenium for 30 h before incubatation with 1 μM H(2)O(2) for 2 h. Lipid peroxidation levels were lower in the control, selenium, selenium + H(2)O(2) groups than in the H(2)O(2) group. GSH-Px activities were higher in the selenium groups than in the H(2)O(2) group. GSH levels were higher in the control, selenium, selenium + H(2)O(2) groups than in the H(2)O(2) group. Cytosolic Ca(2+) release was higher in the H(2)O(2) group than in the control, selenium, selenium + H(2)O(2) groups. Cytosolic Ca(2+) release was lower in the selenium + H(2)O(2) group than in the H(2)O(2). In conclusion, the present study demonstrates that selenium induced protective effects on oxidative stress, [Ca(2+)](c) release and apoptosis in DRG cells. Since selenium deficiency is a common feature of oxidative stress-induced neurological diseases of sensory neurons, our findings are relevant to the etiology of pathology in oxidative stress-induced neurological diseases of the DRG neurons.

Influence of Momordica charantia on oxidative stress-induced perturbations in brain monoamines and plasma corticosterone in albino rats

Objectives:The objective of this study was to evaluate the antistress activity of Momordica charantia (MC) fruit extract on stress-induced changes in albino rats and also to explore attenuating effects of MC on in vitro lipid peroxidation in rat brain.Materials and Methods:In this study, Wistar albino rats (180–200 g) were used. Plasma corticosterone and monoamines—5-hydroxy tryptamine (5-HT), norepinephrine (NE), epinephrine (E) and dopamine (DA) in cortex, hypothalamus and hippocampus regions of brain were determined in animals under different stressful conditions. Ethanolic fruit extract of MC, at doses of 200 and 400 mg/kg, was used. The oxidative stress paradigms used in in vivo models were acute stress (AS) and chronic unpredictable stress (CUS). Panax quinquefolium (PQ) was used as a standard in in vivo models and ascorbic acid was used as a reference standard in the in vitro method.Results:Subjecting the animals to AS (immobilization for 150 min once only) resulted in significant elevation of plasma corticosterone levels and brain monoamine levels. Pretreatment with MC at doses of 200 and 400 mg/kg p.o. significantly countered AS-induced changes and a similar effect was exhibited by PQ at 100 mg/kg p.o. In the CUS regimen (different stressors for 7 days), plasma corticosterone levels were significantly elevated whereas the levels of 5-HT, NE, E, and DA were depleted significantly. Pretreatment with MC (200 and 400 mg/kg) attenuated the CUS-induced changes in the levels of above monoamines in cortex, hypothalamus, and hippocampus regions of brain and plasma corticosterone in a dose-dependent manner. Furthermore, MC extract (1000–5000 μg/mL) exhibited a significant quenching effect on in vitro lipid peroxidation indicating its strong antioxidant activity which was compared with ascorbic acid.Conclusions:This study reveals the antistress activity of MC as it significantly reverted the stress-induced changes, and the activity might be attributed to its antioxidant activity since stress is known to involve several oxidative mechanisms.

Respiratory chain cysteine and methionine usage indicate a causal role for thiyl radicals in aging

The identification of longevity-related structural adaptations in biological macromolecules may yield relevant insights into the molecular mechanisms of aging. In screening fully sequenced animal proteomes for signals associated with longevity, it was found that cysteine depletion in respiratory chain complexes was the by far strongest predictor on the amino acid usage level to co-vary with lifespan. This association was though restricted to aerobic animals, whereas anaerobic animals showed variable cysteine accumulation. By contrast, methionine accumulation, a prominent feature of mitochondrially encoded proteins affording competitive antioxidant protection, was not predictive of longevity, but rather paralleled aerobic metabolic capacity. Hence, the easily oxidized sulfur-containing amino acids cysteine (a thiol) and methionine (a thioether) show doubly diametrical behaviour in two central paradigms of respiratory oxidative stress. From this comparison, it is concluded that only the one–electron oxidation of thiols to thiyl radicals contributes to aging, whereas other forms of sulfur oxidation, especially even-electron oxidation of both thiols and thioethers, are less critically involved, presumably as their consequences may be much more easily repaired. Thiyl radicals may yet act as chain-transfer agents to entail an irreversible intramembrane cross-linking (“plastination”) of some of the a priori most hydrophobic and insoluble proteins known, the respiratory chain complexes.

Nanoparticles: molecular targets and cell signalling

Increasing evidence linking nanoparticles (NPs) with different cellular outcomes necessitate an urgent need for the better understanding of cellular signalling pathways triggered by NPs. Oxidative stress has largely been reported to be implicated in NP-induced toxicity. It could activate a wide variety of cellular events such as cell cycle arrest, apoptosis, inflammation and induction of antioxidant enzymes. These responses occur after the activation of different cellular pathways. In this context, three groups of MAP kinase cascades [ERK (extracellular signal-regulated kinases), p38 mitogen-activated protein kinase and JNK (c-Jun N-terminal kinases)] as well as redox-sensitive transcription factors such as NFκB and Nrf-2 were specially investigated. The ability of NPs to interact with these signalling pathways could partially explain their cytotoxicity. The induction of apoptosis is also closely related to the modulation of signalling pathways induced by NPs. Newly emerged scientific areas of research are the studies on interactions between NPs and biological molecules in body fluids, cellular microenvironment, intracellular components or secreted cellular proteins such as cytokines, growth factors and enzymes and use of engineered NPs to target various signal transduction pathways in cancer therapy. Recently published data present the ability of NPs to interact with membrane receptors leading to a possible aggregation of these receptors. These interactions could lead to a sustained modulation of specific signalling in the target cells or paracrine and even "by-stander" effects of the neighbouring cells or tissues. However, oxidative stress is not sufficient to explain specific mechanisms which could be induced by NPs, and these new findings emphasize the need to revise the paradigm of oxidative stress to explain the effects of NPs.

Traditional reactive carbonyl scavengers do not prevent the carbonylation of brain proteins induced by acute glutathione depletion

This study investigated the effect of reactive carbonyl species (RCS)-trapping agents on the formation of protein carbonyls during depletion of brain glutathione (GSH). To this end, rat brain slices were incubated with the GSH-depletor diethyl maleate in the absence or presence of chemically different RCS scavengers (hydralazine, methoxylamine, aminoguanidine, pyridoxamine, carnosine, taurine and z-histidine hydrazide). Despite their strong reactivity towards the most common RCS, none of the scavengers tested, with the exception of hydralazine, prevented protein carbonylation. These findings suggest that the majority of protein-associated carbonyl groups in this oxidative stress paradigm do not derive from stable lipid peroxidation products like malondialdehyde (MDA), acrolein and 4-hydroxynonenal (4-HNE). This conclusion was confirmed by the observation that the amount of MDA-, acrolein- and 4-HNE-protein adducts does not increase upon GSH depletion. Additional studies revealed that the efficacy of hydralazine at preventing carbonylation was due to its ability to reduce oxidative stress, most likely by inhibiting mitochondrial production of superoxide and/or by scavenging lipid free radicals.

Caspase-mediated protein kinase C-δ cleavage is necessary for apoptosis of vascular smooth muscle cells

Apoptotic death of vascular smooth muscle cells (SMCs) is a prominent feature of blood vessel remodeling and various vascular diseases. We have previously shown that protein kinase C-delta (PKC-delta) plays a critical role in SMC apoptosis. In this study, we tested the importance of PKC-delta proteolytic cleavage and tyrosine phosphorylation within the apoptosis pathway. Using hydrogen peroxide as a paradigm for oxidative stress, we showed that proteolytic cleavage of PKC-delta occurred in SMCs that underwent apoptosis, while tyrosine phosphorylation was detected only in necrotic cells. Furthermore, using a peptide (z-DIPD-fmk) that mimics the caspase-3 binding motif within the linker region of PKC-delta, we were able to prevent the cleavage of PKC-delta, as well as apoptosis. Inhibition of PKC-delta with rottlerin or small-interfering RNA diminished caspase-3 cleavage, caspase-3 activity, cleavage of poly (ADP-ribose) polymerase, cleavage of PKC-delta, and DNA fragmentation, confirming the previously reported role of PKC-delta in initiation of apoptosis. In contrast, z-DIPD-fmk markedly diminished caspase-3 activity, cleavage of PKC-delta, and DNA fragmentation without affecting cleavage of caspase-3 and poly (ADP-ribose) polymerase. Taken together, our data suggest that caspase-3-mediated PKC-delta cleavage underlies SMC apoptosis induced by oxidative stress, and that PKC-delta acts both upstream and downstream of caspase-3.

Assessing Steady-state Fluorescence and PRI from Hyperspectral Proximal Sensing as Early Indicators of Plant Stress: The Case of Ozone Exposure.

High spectral resolution spectrometers were used to detect optical signals ofongoing plant stress in potted white clover canopies subjected to ozone fumigation. Thecase of ozone stress is used in this manuscript as a paradigm of oxidative stress. Steadystatefluorescence (Fs) and the Photochemical Reflectance Index (PRI) were investigatedas advanced hyperspectral remote sensing techniques able to sense variations in the excessenergy dissipation pathways occurring when photosynthesis declines in plants exposed to astress agent. Fs and PRI were monitored in control and ozone fumigated canopies during a21-day experiment together with the traditional Normalized Difference Vegetation Index(NDVI) and physiological measurements commonly employed by physiologists to describestress development (i.e. net CO₂ assimilation, active fluorimetry, chlorophyll concentrationand visible injuries). It is shown that remote detection of an ongoing stress through Fs andPRI can be achieved in an early phase, characterized by the decline of photosynthesis. Onthe contrary, NDVI was able to detect the stress only when damage occurred. These resultsopen up new possibilities for assessment of plant stress by means of hyperspectral remotesensing.

Comparison of the Biological Activity Between Ultrafine and Fine Titanium Dioxide Particles in RAW 264.7 Cells Associated with Oxidative Stress

Ultrafine or fine titanium dioxide (TiO2) particles are widely used in the production of white pigments, for sunscreens, and in cleanup techniques. However, currently knowledge is deficient concerning cellular responses to these particles. The study evaluated and compared the biological activity of ultrafine and fine TiO2 particles in RAW 264.7 macrophages according to an oxidative stress paradigm. In vitro exposure of macrophages to ultrafine or fine TiO2 in the range of 0.5–200 μg/ml did not significantly alter cell viability. However, ultrafine TiO2 enhanced intracellular generation of reactive oxygen species (ROS) to a greater extent than fine TiO2 at each exposure concentration. Ultrafine TiO2 induced ERK1/2 activation in a concentration-dependent manner, while the fine TiO2-induced changes were minimal. Phosphorylation of ERK1/2 occurred following 10 min exposure to higher concentrations of ultrafine TiO2 (≥25 μg/ml). Similarly, ultrafine TiO2 exposure significantly enhanced tumor necrosis factor (TNF)-α and macrophage inflammatory protein (MIP)-2 secretion in a concentration-dependent manner, and its potency was higher than fine TiO2. These findings suggest that when exposure concentration is based upon equivalent mass, ultrafine TiO2 exerts greater biological activity as measured by ROS generation, ERK 1/2 activation, and proinflammatory mediator secretion in RAW 264.7 macrophages than fine TiO2.

Protection against Oxidant-Induced Apoptosis by Exogenous Glutathione in Leber Hereditary Optic Neuropathy Cybrids

To use different paradigms of oxidative and metabolic stress in a cellular model of Leber hereditary optic neuropathy (LHON), with the aim of evaluating the efficacy of potentially therapeutic molecules for the treatment of this disease. Cybrids bearing one of the three most common LHON pathogenic mutations (11778/ND4, 3460/ND1, 14484/ND6) were incubated with two compounds known to induce oxidative injury, tert-butyl hydroperoxide (t-BH) and rotenone. To mimic metabolic stress, cells were incubated in a glucose-free medium containing galactose. Cell viability was determined using the MTT assay. To identify the apoptotic type of cell death, nuclear morphology was examined after cell loading with Hoechst. Cellular glutathione (GSH), and oxidized glutathione (GSSG) levels were measured enzymatically. Incubation with t-BH caused apoptotic cell death of control and LHON cybrids, whereas only LHON cybrids were damaged by rotenone concentrations up to 2.5 muM. Both types of stress caused a marked imbalance in the glutathione levels, but an increase in the GSSG/GSH+GSSG ratio was detected only after rotenone treatment. The efficacy of several antioxidant and antiapoptotic compounds was then assessed in cells exposed to these two oxidative paradigms. Only exogenous GSH remarkably protected the t-BH- and rotenone-treated cybrids from cell death. In contrast, GSH was unable to increase the viability of cybrids exposed to metabolic stress. These results suggest that GSH is an effective antioxidant compound to be tested as a potential treatment for LHON.

Mitochondrial oxidative and structural damage in ischemia-reperfusion in human myocardium. Current knowledge and future directions

The sequence of events in heart ischemia-reperfusion has been clearly documented in experimental animal models but not in cardiac surgery patients. The evidence in human studies had not been gathered in a systematic and comprehensive fashion, so as to provide an encompassing picture of the phenomenon. This limits our ability to devise appropriate strategies for optimal perioperative myocardial protection. We present here a review or our experience in myocardial ischemia-reperfusion in a historical perspective. From our previous studies we conclude that, although several issues still remain unsolved, there is no doubt that oxygen-free radicals are important contributors to myocardial injury during the reperfusion period of coronary artery bypass surgery. Yet, in spite of this wealth of information, both clinical and experimental, subsequent clinical trials conducted over the last several years with a variety of antioxidant strategies have been largely disappointing. Therefore, the whole paradigm of oxidative stress in cardiac injury needs to be re-evaluated. In this regard, differences between past and current knowledge are discussed, and future directions are traced. We concluded that patients subjected to elective bypass surgery undergo oxidative stress upon reperfusion after cardioplegic arrest; the magnitude of the phenomenon, however, is at present small and may not justify widespread antioxidant therapy.

Comparison of the Abilities of Ambient and Manufactured Nanoparticles To Induce Cellular Toxicity According to an Oxidative Stress Paradigm

Nanomaterial properties differ from those bulk materials of the same composition, allowing them to execute novel activities. A possible downside of these capabilities is harmful interactions with biological systems, with the potential to generate toxicity. An approach to assess the safety of nanomaterials is urgently required. We compared the cellular effects of ambient ultrafine particles with manufactured titanium dioxide (TiO2), carbon black, fullerol, and polystyrene (PS) nanoparticles (NPs). The study was conducted in a phagocytic cell line (RAW 264.7) that is representative of a lung target for NPs. Physicochemical characterization of the NPs showed a dramatic change in their state of aggregation, dispersibility, and charge during transfer from a buffered aqueous solution to cell culture medium. Particles differed with respect to cellular uptake, subcellular localization, and ability to catalyze the production of reactive oxygen species (ROS) under biotic and abiotic conditions. Spontaneous ROS production was compared by using an ROS quencher (furfuryl alcohol) as well as an NADPH peroxidase bioelectrode platform. Among the particles tested, ambient ultrafine particles (UFPs) and cationic PS nanospheres were capable of inducing cellular ROS production, GSH depletion, and toxic oxidative stress. This toxicity involves mitochondrial injury through increased calcium uptake and structural organellar damage. Although active under abiotic conditions, TiO2 and fullerol did not induce toxic oxidative stress. While increased TNF-alpha production could be seen to accompany UFP-induced oxidant injury, cationic PS nanospheres induced mitochondrial damage and cell death without inflammation. In summary, we demonstrate that ROS generation and oxidative stress are a valid test paradigm to compare NP toxicity. Although not all materials have electronic configurations or surface properties to allow spontaneous ROS generation, particle interactions with cellular components are capable of generating oxidative stress.

TRPM2: a calcium influx pathway regulated by oxidative stress and the novel second messenger ADP-ribose

A unique functional property within the transient receptor potential (TRP) family of cation channels is the gating of TRP (melastatin) 2 (TRPM2) channels by ADP-ribose (ADPR). ADPR binds to the intracellular C-terminal tail of TRPM2, a domain that shows homology to enzymes with pyrophosphatase activity. Cytosolic Ca(2+) enhances TRPM2 gating by ADPR; ADPR and Ca(2+) in concert may be an important messenger system mediating Ca(2+) influx. Other stimuli of TRPM2 include NAD and H(2)O(2) and cyclic ADPR, which may act synergistically with ADPR. H(2)O(2), an experimental paradigm of oxidative stress, may also induce the formation of ADPR in the nucleus or mitochondria. In this review, we summarize the gating properties of TRPM2 and the proposed pathways of channel activation in vivo. TRPM2 is likely to be a key player in several signalling pathways, mediating cell death in response to oxidative stress or in reperfusion injury. Moreover, it plays a decisive role in experimentally induced diabetes mellitus and in the activation of leukocytes.

Impact of oxidative stress on neuronal survival.

1. Reactive oxygen species and oxidative state are slowly gaining acceptance in having a physiological relevance rather than just being the culprits in pathophysiological processes. The control of the redox environment of the cell provides for additional regulation in relation to signal transduction pathways. Conversely, aberrant regulation of oxidative state manifesting as oxidative stress can predispose a cell to adverse outcome. 2. The phosphatidylinositol 3-kinase/akt pathway is one such pathway that is partially regulated via oxidative state and, in an oxidative stress paradigm such as ischaemic-reperfusion injury, may be inactivated, which can lead to exacerbation of cell death. 3. Activation of nuclear factor (NF)-kappaB has been associated with oxidative stress. The role of NF-kappaB in neuronal cell death is widely debated, with major studies highlighting both a pro- and anti-apoptotic role for NF-kappaB, with the outcome being region, stimulus, dose and duration specific. 4. Oxidative state plays a key role in the regulation and control of numerous signal transduction pathways in the cell. Elucidating the mechanisms behind oxidative stress-mediated neuronal cell death is important in identifying potential putative targets for the treatment of diseases such as stroke.

Transient oxidative stress in SH‐SY5Y human neuroblastoma cells results in caspase dependent and independent cell death and tau proteolysis

The effects of an oxidative insult on cell survival and tau metabolism were investigated in human neuroblastoma SH-SY5Y cells. In this treatment paradigm cells were exposed to the membrane permeant oxidant tert-butylhydroperoxide (tBHP) for 40 min, returned to fresh media and cell survival/death was monitored during the post-treatment period. Cell viability decreased significantly by 6 hr after tBHP exposure, and by 24 hr lactate dehydrogenase (LDH) release was 40.1 +/- 8.8% in tBHP treated cells compared to 8.1 +/- 4.7% in control cells. This oxidative stress paradigm also resulted in significant activation of caspase-3 by 2 hr post-treatment and nuclear apoptotic morphology. Furthermore, tBHP treatment also resulted in delayed tau proteolysis that was first evident 2 hr post-treatment. Treatment of the cells with the general caspase inhibitor Boc-Asp(OMe)-Fluoromethylketone (BAF) completely inhibited caspase-3 activation in response to tBHP, and delayed, but did not prevent cell death. BAF treatment also decreased tau proteolysis. In vitro, recombinant tau was readily proteolyzed by active recombinant caspase-3 into a stable breakdown product. Further tau in the cell lysates was cleaved by active recombinant caspase-3 at a rate, and to an extent similar to that observed for the well-established caspase-3 substrate poly(ADP-ribose)polymerase (PARP). These results suggest that oxidative stress-induced cell death occurs through both caspase-dependent and-independent pathways, and that tau is likely an in situ substrate of caspase-3.

Transient oxidative stress in SH-SY5Y human neuroblastoma cells results in caspase dependent and independent cell death and tau proteolysis

The effects of an oxidative insult on cell survival and tau metabolism were investigated in human neuroblastoma SH-SY5Y cells. In this treatment paradigm cells were exposed to the membrane permeant oxidant tert-butylhydroperoxide (tBHP) for 40 min, returned to fresh media and cell survival/death was monitored during the post-treatment period. Cell viability decreased significantly by 6 hr after tBHP exposure, and by 24 hr lactate dehydrogenase (LDH) release was 40.1 +/- 8.8% in tBHP treated cells compared to 8.1 +/- 4.7% in control cells. This oxidative stress paradigm also resulted in significant activation of caspase-3 by 2 hr post-treatment and nuclear apoptotic morphology. Furthermore, tBHP treatment also resulted in delayed tau proteolysis that was first evident 2 hr post-treatment. Treatment of the cells with the general caspase inhibitor Boc-Asp(OMe)-Fluoromethylketone (BAF) completely inhibited caspase-3 activation in response to tBHP, and delayed, but did not prevent cell death. BAF treatment also decreased tau proteolysis. In vitro, recombinant tau was readily proteolyzed by active recombinant caspase-3 into a stable breakdown product. Further tau in the cell lysates was cleaved by active recombinant caspase-3 at a rate, and to an extent similar to that observed for the well-established caspase-3 substrate poly(ADP-ribose)polymerase (PARP). These results suggest that oxidative stress-induced cell death occurs through both caspase-dependent and-independent pathways, and that tau is likely an in situ substrate of caspase-3.