4-hydroxynonenal induces mitochondrial oxidative stress, apoptosis and expression of glutathione S-transferase A4-4 and cytochrome P450 2E1 in PC12 cells

An excessive and sustained increase in reactive oxygen species (ROS) production and oxidative stress have been implicated in the pathogenesis of many diseases. In the present study, we have demonstrated that 4-hydroxynonenal (4-HNE), a product of lipid peroxidation, alters glutathione (GSH) pools and induces oxidative stress in PC12 cells in culture. This increase was accompanied by alterations in subcellular ROS and glutathione (GSH) metabolisms. The GSH homeostasis was affected as both mitochondrial and extramitochondrial GSH levels, GSH peroxidase and glutathione reductase activities were inhibited and glutathione S-transferase (GST) activity was increased after 4-HNE treatment. A concentration- and time-dependent increase in cytochrome P450 2E1 (CYP 2E1) activity in the mitochondria and postmitochondrial supernatant was also observed. 4-HNE-induced oxidative stress also caused an increase in the expression of GSTA4-4, CYP2E1 and Hsp70 proteins in the mitochondria. Increased oxidative stress in PC12 cells initiated apoptosis as indicated by the release of mitochondrial cytochrome c, activation of poly-(ADP-ribose) polymerase (PARP), DNA fragmentation and decreased expression of antiapoptotic Bcl-2 proteins. Mitochondrial respiratory and redox functions also appeared to be affected markedly by 4-HNE treatment. These results suggest that HNE-induced oxidative stress and apoptosis might be associated with altered mitochondrial functions and a compromised GSH metabolism and ROS clearance.     

Increased mitochondrial stress and modulation of mitochondrial respiratory enzyme activities in acetaminophen-induced toxicity in mouse macrophage cells

Overdose of acetaminophen (APAP) causes tissue injury particularly in the liver. However, the precise mechanism of APAP toxicity is not clear. Glutathione (GSH) depletion and oxidative stress are believed to be the main cause of APAP toxicity. The role of macrophages in APAP-induced tissue injury is controversial. Using mouse macrophage J774.2 cells, we recently demonstrated that like in animal models, APAP reduces GSH pool and alters GSH metabolism by increasing the production of reactive oxygen species (ROS). In the present study, we show that APAP-induced cytotoxicity and apoptosis in macrophages are associated with increased mitochondrial metabolic and oxidative stress, alterations in the mitochondrial membrane potential and activities of the respiratory enzyme complexes. APAP treatment also altered ROS/NO production and inhibited the expression of COX-2 and iNOS in LPS-stimulated macrophages. Electron microscopic studies also confirmed morphological changes associated with apoptosis at the lower dose of APAP, while at the higher dose late apoptosis/necrotic changes were visible. These results suggest that mitochondrial metabolic and oxidative stress are the main causes of cytotoxicity and cell death in APAP treated macrophages. The study may have long term implications to better understand the role of macrophages in the toxicology and pharmacology of APAP.     

Green tea polyphenol epigallocatechin-3-gallate differentially modulates oxidative stress in PC12 cell compartments

Tea polyphenols have been reported to be potent antioxidants and beneficial in oxidative stress related diseases. Prooxidant effects of tea polyphenols have also been reported in cell culture systems. In the present study, we have studied oxidative stress in the subcellular compartments of PC12 cells after treatment with different concentrations of the green tea polyphenol, epigallocatechin-3-gallate (EGCG). We have demonstrated that EGCG has differentially affected the production of reactive oxygen species (ROS), glutathione (GSH) metabolism and cytochrome P450 2E1 activity in the different subcellular compartments in PC12 cells. Our results have shown that although the cell survival was not inhibited by EGCG, there was, however, an increased DNA breakdown and activation of apoptotic markers, caspase 3 and poly- (ADP-ribose) polymerase (PARP) at higher concentrations of EGCG treatment. Our results suggest that the differential effects of EGCG might be related to the alterations in oxidative stress, GSH pools and CYP2E1 activity in different cellular compartments. These results may have implications in determining the chemopreventive therapeutic use of tea polyphenols in vivo.     

Reactive oxygen species regulated mitochondria-mediated apoptosis in PC12 cells exposed to chlorpyrifos

Reactive oxidative species (ROS) generated by environmental toxicants including pesticides could be one of the factors underlying the neuronal cell damage in neurodegenerative diseases. In this study we found that chlorpyrifos (CPF) induced apoptosis in dopaminergic neuronal components of PC12 cells as demonstrated by the activation of caspases and nuclear condensation. Furthermore, CPF also reduced the tyrosine hydroxylase-positive immunoreactivity in substantia nigra of the rat. In addition, CPF induced inhibition of mitochondrial complex I activity. Importantly, N-acetyl cysteine (NAC) treatment effectively blocked apoptosis via the caspase-9 and caspase-3 pathways while NAC attenuated the inhibition of mitochondrial complex I activity as well as the oxidative metabolism of dopamine (DA). These results demonstrated that CPF-induced apoptosis was involved in mitochondrial dysfunction through the production of ROS. In the response of cellular antioxidant systems to CPF, we found that CPF treatment increased HO-1 expression while the expression of CuZnSOD and MnSOD was reduced. In addition, we found that CPF treatment activated MAPK pathways, including ERK 1/2, the JNK, and the p38 MAP kinase in a time-dependent manner. NAC treatment abolished MAPK phosphorylation caused by CPF, indicating that ROS are upstream signals of MAPK. Interestingly, MAPK inhibitors abolished cytotoxicity and reduced ROS generation by CPF treatment. Our results demonstrate that CPF induced neuronal cell death in part through MAPK activation via ROS generation, suggesting its potential to generate oxidative stress via mitochondrial damage and its involvement in oxidative stress-related neurodegenerative disease.     

Modulation of apoptosis and improved redox metabolism with the use of a new antioxidant formula

Oxidative stress is involved in the pathogenesis of a wide spectrum of diseases, implicating that strategies directed at counterbalancing oxidative processes could have a role in clinical medicine. There is also an evidence that oxidative stress acts as a major determinant of apoptotic cell death. Many studies have reported favourable effects of antioxidant formulas on several parameters of the oxidant-antioxidant balance, but none of them has focused whether antioxidant formulas could modulate apoptosis. We investigated in 20 healthy individuals the effect of supplementation with a formula containing alpha-tocopherol, alpha-lipoic acid, coenzyme Q(10), carnitines, and selenomethionine, on plasma oxidant status and peroxide levels, erythrocyte antioxidant enzymes, lymphocyte apoptosis, and generation of ROS at the mitochondrial level. Control subjects received only carnitines or an incomplete formula with alpha-tocopherol, alpha-lipoic acid, coenzyme Q(10), and selenomethionine. Supplementation with the complete formula resulted in a significant increase in the plasma antioxidant status that was mirrored by a decrease in blood peroxide levels and a reduced generation of ROS at the mitochondrial level. This was associated with a significant decrease in the frequency of peripheral blood lymphocytes, with either CD4 or CD8 phenotype, undergoing apoptosis. Less consistent results were found when either incomplete formula was used. Our study suggests that supplementation with antioxidant formulas can modulate the process of apoptosis under in vivo conditions. The clinical potential of this strategy in the treatment of diseases with an elevated commitment to apoptosis should be explored.     

In vitro protection of reactive oxygen species-induced degradation of lipids, proteins and 2-deoxyribose by tea catechins.

Both the anti- and pro-oxidant effects of tea catechins, have been implicated in the alterations of cellular functions which determine their chemoprotective and therapeutic potentials in toxicity and diseases. Here, we have studied the protective mechanism (s) of three main green tea catechins namely, epicatechin (EC), epicatechin gallate (ECG) and epigallocatechin gallate (EGCG) on free radical induced oxidative degradation of membrane lipids and proteins under in vitro conditions using isolated cell free fractions from rat liver. In addition, we have also studied the effects of the tea catechins on 2-deoxyribose degradation in the presence of Fenton and Haber-Weiss oxidants. Glutathione S-transferase and cytochrome P450 2E1 activities and lipid peroxidation were found to be markedly inhibited by tea catechins. These catechins also inhibited the reactive oxygen species formation and oxidative carbonylation of subcellular proteins induced by a physiological oxidant, 4-hydroxynonenal. EGCG and the other catechins showed a time and concentration-dependent effects on the degradation of 2-deoxyribose in the presence of Fenton oxidants. Our results indicate that tea catechins prevent molecular degradation in oxidative stress conditions by directly altering the subcellular ROS production, glutathione metabolism and cytochrome P450 2E1 activity. These results may have implications in determining the chemotherapeutic use of tea catechins in oxidative stress related diseases.     

A rapid and transient ROS generation by cadmium triggers apoptosis via caspase-dependent pathway in HepG2 cells and this is inhibited through N-acetylcysteine-mediated catalase upregulation

Although reactive oxygen species (ROS) have been implicated in cadmium (Cd)-induced hepatotoxicity, the role of ROS in this pathway remains unclear. Therefore, we attempted to determine the molecular mechanisms relevant to Cd-induced cell death in HepG2 cells. Cd was found to induce apoptosis in the HepG2 cells in a time- and dose-dependent fashion, as confirmed by DNA fragmentation analysis and TUNEL staining. In the early stages, both rapid and transient ROS generation triggered apoptosis via Fas activation and subsequent caspase-8-dependent Bid cleavage, as well as by calpain-mediated mitochondrial Bax cleavage. The timing of Bid activation was coincided with the timing at which the mitochondrial transmembrane potential (MMP) collapsed as well as the cytochrome c (Cyt c) released into the cytosol. Furthermore, mitochondrial permeability transition (MPT) pore inhibitors, such as cyclosporin A (CsA) and bongkrekic acid (BA), did not block Cd-induced ROS generation, MMP collapse and Cyt c release. N-acetylcysteine (NAC) pretreatment resulted in the complete inhibition of the Cd-induced apoptosis via catalase upregulation and subsequent Fas downregulation. NAC treatment also completely blocked the Cd-induced intracellular ROS generation, MMP collapse and Cyt c release, indicating that Cd-induced mitochondrial dysfunction may be regulated indirectly by ROS-mediated signaling pathway. Taken together, a rapid and transient ROS generation by Cd triggers apoptosis via caspase-dependent pathway and subsequent mitochondrial pathway. NAC inhibits Cd-induced apoptosis through the blocking of ROS generation as well as the catalase upregulation.     

Role of oxidative stress and apoptosis in cadmium induced thymic atrophy and splenomegaly in mice

Cadmium immunotoxicity in rodents is primarily characterized by marked thymic damage and splenomegaly. To understand the toxicity of Cd on lymphoid cells in vivo, a single dose of Cd as CdCl2 (1.8 mg/kg, i.p.) was administered to male BALB/c mice and cytotoxicity (MTT assay), oxidative stress indicators (glutathione, reactive oxygen species) and apoptotic markers (mitochondrial membrane potential, caspase-3 activity, phosphatidylserine externalization, apoptotic DNA, intranucleosomal DNA fragmentation) were assessed in thymic and splenic single cell suspensions, at various time intervals. Lowering of body weight gain and cellularity and a loss in cell viability was seen in the Cd treated mice. The earliest significant increase in ROS at 18 h, followed by mitochondrial membrane depolarization, caspase-3 activation and GSH depletion at 24h in spleen and later at 48 h in thymus, strongly implicate the possible involvement of ROS. A pronounced inhibition of cell proliferative response at 48 h and 72 h may also be linked to Cd induced apoptosis. The morphological alterations including thymic cortical cell depletion and an increase in red pulp with diminished white pulp in spleen were observed at 48 h and beyond. The splenic cells appeared more susceptible than thymus cells to the adverse effects of Cd. The present study, therefore, demonstrates potentiation of oxidative stress followed by mitochondrial-caspase dependent apoptotic pathway. This may, in part, be responsible for causing suppression of cell proliferative response, thymic atrophy and splenomegaly.     

Casiopeína IIgly-induced oxidative stress and mitochondrial dysfunction in human lung cancer A549 and H157 cells

Casiopeínas are a series of mixed chelate copper complexes that are being evaluated as anticancer agents. Their effects in the cell include oxidative damage and mitochondrial dysfunction, yet the molecular mechanisms leading to such effects remain unclear. We tested whether [Cu(4,7-dimethyl-phenanthroline)(glycinate)]NO₃ (Casiopeína IIgly or Cas IIgly) could alter cellular glutathione (GSH) levels by redox cycling with GSH to generate ROS and cellular oxidative stress. Cas IIgly induced a dramatic drop in intracellular levels of GSH in human lung cancer H157 and A549 cells, and is able to use GSH as source of electrons to catalyze the Fenton reaction. In both cell lines, the toxicity of Cas IIgly (2.5–5 μM) was potentiated by the GSH synthesis inhibitor l-buthionine sulfoximine (BSO) and diminished by the catalytic antioxidant manganese(III) meso-tetrakis(N,N′-diethylimidazolium-2-yl)porphyrin (MnTDE-1,3-IP⁵⁺), thus supporting an important role for oxidative stress. Cas IIgly also caused an over-production of reactive oxygen species (ROS) in the mitochondria and a depolarization of the mitochondrial membrane. Moreover, Cas IIgly produced mitochondrial DNA damage that resulted in an imbalance of the expression of the apoproteins of the mitochondrial respiratory chain, which also can contribute to increased ROS production. These results suggest that Cas IIgly initiates multiple possible sources of ROS over-production leading to mitochondrial dysfunction and cell death.     

Cytotoxic effects of incense particles in relation to oxidative stress,the cell cycle and F-actin assembly

Epidemiological studies have suggested that combustion-derived smoke, such as that produced during incense burning, is a deleterious air pollutant. It is capable of initiating oxidative stress and mutation; however, the related apoptotic processes remain unclear. In order to elucidate the biological mechanisms of reactive oxygen species (ROS)-induced respiratory toxicology, alveolar epithelial A549 cells were exposed to incense particulate matter (PM), with and without antioxidant N-acetyl-l-cysteine (NAC). The cross-linking associations between oxidative capacity, cell cycle events, actin cytoskeletal dynamics and intracellular calcium signals were investigated. An incense PM suspension caused significant oxidative stress in A549 cells, as shown by inhibition of the cell cycle at G1 and G2/M check-points, and the induction of apoptosis at Sub-G1. At the same time, alterations in the F-actin filamentous assemblies were observed. The levels of intracellular Ca²⁺ were increased after incense PM exposure. Antioxidant NAC treatment revealed that oxidative stress and F-actin remodelling was significantly mitigated. This suggests that ROS accumulation could alter cell cycle regulation and anomalous remodelling of the cortical cytoskeleton that allowed impaired cells to enter into apoptosis. This study has elucidated the integral patho-physiological interactions of incense PM and the potential mechanisms for the development of ROS-driven respiratory impairment.     

Antioxidant and pro-oxidant properties of pyrroloquinoline quinone (PQQ): implications for its function in biological systems

Pyrroloquinoline quinone (PQQ) is a novel redox cofactor recently found in human milk. It has been reported to function as an essential nutrient, antioxidant and redox modulator in cell culture experiments and in animal models of human diseases. As mitochondria are particularly susceptible to oxidative damage we studied the antioxidant properties of PQQ in isolated rat liver mitochondria. PQQ was an effective antioxidant protecting mitochondria against oxidative stress-induced lipid peroxidation, protein carbonyl formation and inactivation of the mitochondrial respiratory chain. In contrast, PQQ caused extensive cell death to cells in culture. This surprising effect was inhibited by catalase, and was shown to be due to the generation of hydrogen peroxide during the autoxidation of PQQ in culture medium. We conclude that the reactivities of PQQ are dependent on its environment and that it can act as an antioxidant or a pro-oxidant in different biological systems.     

The course of CCl4 induced hepatotoxicity is altered in mGSTA4-4 null (-/-) mice

Glutathione S-transferases (GSTs) play a key role in cellular detoxification of environmental toxicants through their conjugation to glutathione (GSH). Recent studies have shown that the alpha-class GSTs also provide protection against oxidative stress and lipid peroxidation (LPO). GSTA4-4 is a member of a sub group of the alpha-class GSTs. It has been shown to metabolize 4-hydroxynonenal (4-HNE) with high catalytic efficiency through its conjugation to glutathione (GSH) and has been suggested to be a major component of cellular defense against toxic electrophiles such as 4-HNE generated during LPO. Since the hepatotoxicity of carbon tetrachloride (CCl(4)) has been suggested to be due to the generation of free radicals leading to membrane LPO, the present studies were designed to compare hepatotoxicity of CCl(4) in GSTA4-4 null (-/-) and wild type (+/+) mice. The results show that administration of a single dose of CCl(4) (1 ml/kg i.p.) resulted in time dependent hepatotoxicity in both -/- and +/+ mice; the extent of cellular damage by serum enzymes suggests that progression was more rapid in -/- mice, although injury was similar by 24 h. Histopathologic examination showed similar degrees of centrilobular necrosis by 24 h but much greater surrounding degenerative change, including cellular swelling, disarray, and vacuolization, in the liver of -/- mice. As expected -/- mice did not show any expression of mGSTA4-4; after CCl(4) a compensatory increase in the activities of total GST activity was noted at 24 h. Major alterations in other antioxidant enzymes was not observed. 4-HNE levels in the liver of -/- mice were about four-fold higher than in +/+ mice, suggesting a positive correlation between 4-HNE levels and the altered course of CCl(4) hepatotoxicity. These studies suggest that GSTA4-4 is an important component during the early stages (1-6 h) of cellular defense against oxidative stress and LPO although, it is not effective in protecting against the ultimate degree of overall cell injury.     

Glycyrrhizic acid modulates t-BHP induced apoptosis in primary rat hepatocytes

Glycyrrhizic acid (GA) is the main bioactive ingredient of licorice (Glycyrrhiza glabra). The object of this study was to evaluate the protective effects of GA on tert-butyl hydroperoxide (t-BHP) induced oxidative injury leading to apoptosis in cultured primary rat hepatocytes. Throughout the study silymarin was used as positive control. Molecular mechanisms involved in apoptotic pathways induced in hepatocytes by t-BHP at 250 μM were explored in detail. DNA fragmentation, activation of caspases and cytochrome c release were demonstrated. In addition, changes in the mitochondrial membrane potential and ROS generation were detected confirming involvement of mitochondrial pathway. Pre-treatment with GA (4 μg) protected the hepatocytes against t-BHP induced oxidative injury and the results were comparable to the pre-treatment with positive control, i.e. silymarin. The protective potential against cell death was achieved mainly by preventing intracellular GSH depletion, decrease in ROS formation as well as inhibition of mitochondrial membrane depolarization. GA was found to modulate critical end points of oxidative stress induced apoptosis and could be beneficial against liver diseases where oxidative stress is known to play a crucial role.     

Cadmium induces apoptotic cell death in WI 38 cells via caspase-dependent Bid cleavage and calpain-mediated mitochondrial Bax cleavage by Bcl-2-independent pathway

Previous reports have demonstrated that cadmium (Cd) may induce cell death via apoptosis, but the mechanism responsible for cellular death is not clear. In this study, we investigated the signaling pathways implicated in Cd-induced apoptosis in lung epithelial fibroblast (WI 38) cells. Apoptotic features were observed using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, propidium iodide staining and DNA laddering. A treatment of cadmium caused the caspase-8-dependent Bid cleavage, the release of cytochrome c (Cyt c), activation of caspase-9 and -3, and PARP cleavage. A caspase-8 specific inhibitor prevented the Bid cleavage, caspase-3 activation and cell death. Alternatively, we observed that full-length Bax was cleaved into 18-kDa fragment (p18/Bax); this was initiated after 12 h and by 36 h the full-length Bax protein was totally cleaved to the p18/Bax, which caused a drastic release of Cyt c from mitochondria. The p18/Bax was detected exclusively in the mitochondrial fraction, and it originated from mitochondrial full-length Bax, but not from the cytosol full-length Bax. Cd also induced the activation of the mitochondrial 30-kDa small subunit of calpain that was preceded by Bax cleavage. Cd induced the upregulation of Bcl-2 and the degradation of p53 protein. N-acetyl cysteine effectively inhibited the Cd-induced DeltaPsim reduction, indicating ROS acts upstream of mitochondrial membrane depolarization. Taken together, our results suggest that Cd-induced apoptosis was thought to be mediated at least two pathways; caspase-dependent Bid cleavage, and the other is calpain-mediated mitochondrial Bax cleavage. Moreover, we found that the function of Bid and Bax was not dependent of Bcl-2, and that ROS can also contribute in the Cd-induced cell death.     

Role of reactive oxygen species, glutathione and NF-kappaB in apoptosis induced by 3,4-methylenedioxymethamphetamine ("Ecstasy") on hepatic stellate cells

"Ecstasy" (3,4-methylenedioxymethamphetamine, MDMA), is a derivative of amphetamine with hepatotoxic effects that has been shown to induce apoptosis of cultured liver cells. In the present work, we studied the role played by oxidative stress in the apoptotic response caused by MDMA on a cell line of hepatic stellate cells (HSC). MDMA-treatment provoked oxidative stress determined as reactive oxygen species (ROS) accumulation and decrease of intracellular reduced glutathione levels. Pre-treatment with the antioxidant pyrrolidine dithiocarbamate blocked ROS production but did not prevent MDMA-induced apoptosis of HSC. The pro-oxidant menadione induced in HSC ROS production and apoptosis that were prevented by pyrrolidine dithiocarbamate, showing HSC to be susceptible to oxidative stress-induced apoptosis. Addition of exogenous GSH or its precursor NAC potentiated the apoptotic action of MDMA but blocked apoptosis induced by menadione. Pre-treatment of HSC with the cytochrome P450 inhibitor quinine diminished the extent of apoptosis caused by MDMA, suggesting the involvement of a metabolic derivative of MDMA on its apoptotic effect. Nuclear factor NF-kappaB was activated by MDMA in a oxidative stress independent fashion and played a protective role in the apoptotic response, since inhibition of NF-kappaB by treatment with parthenolide or by viral infection with a dominant-negative form of NIK (Ad5dnNIK) resulted in an increase of MDMA-induced cell death. In summary, MDMA-induced apoptosis of HSC is accompanied, but not caused by oxidative stress; a metabolic derivative of the drug is responsible for the apoptotic effect of MDMA, which is partially blocked by NF-kappaB activation.     

Mitochondria,reactive oxygen species and cadmium toxicity in the kidney

The heavy metal cadmium accumulates in kidney cells, particularly those of the proximal tubular epithelium, and the damage this causes is associated with development of chronic kidney disease. One of the causative mechanisms of chronic kidney disease is thought to be oxidative stress. Cadmium induces oxidative stress, but the molecular mechanisms involved in the cell damage from oxidative stress in cadmium-induced chronic kidney disease are not well understood. Mitochondrial damage is likely, given that dysfunctional mitochondria are central to the formation of excess reactive oxygen species (ROS), and are known key intracellular targets for cadmium. Normally, ROS are balanced by natural anti-oxidant enzymes. When mitochondria become dysfunctional, for example, through long term exposure to environmental toxicants like cadmium, they produce less cell energy and more ROS. The imbalance between these ROS and the natural anti-oxidants creates the condition of oxidative stress. The outcomes of mitochondrial injury are manyfold: injured mitochondria perpetuate oxidative stress; the loss of mitochondrial membrane potential causes release of cytochrome-c and activation of caspase pathways that lead to apoptotic deletion of renal cells; and attempts by cells to remove dysfunctional mitochondria through autophagy lead to “autophagic cell death” or apoptosis. Three pathways of mitochondrial regulation (upstream signalling pathways, direct mitochondrial targeting, and downstream cell death effector pathways) are therefore all promising targets for effective anti-oxidant treatment of cadmium toxicity in the kidney.     

Mitochondrial state 3 to 4 respiration transition during Fas-mediated apoptosis controls cellular redox balance and rate of cell death

The role of reactive oxygen species (ROS) production in death receptor-mediated apoptosis is ill defined. We show that ROS levels play a novel role in moderating the rate of cell death in Fas-dependent apoptosis. Treatment of Jurkat T cells with oligomycin (ATP-synthase inhibitor) or FCCP (mitochondrial uncoupler) and Fas activating antibody (CH11), facilitated rapid cell death. ATP levels, DEVDase activity and cytochrome c release were not account for the synergistic killing effect. However, a decrease in cellular ROS production was associated with CH11 treatment and combinations of CH11 with oligomycin or FCCP further inhibited cellular ROS levels. Thus, decreased ROS production is correlated with accelerated cell death. A transition from state 3 to state 4 mitochondrial respiration following apoptotic stimuli accounted for an attenuated membrane potential and as a results mitochondria-derived ROS production capacity diminished. Similar observations were demonstrated in isolated rat liver mitochondria. Transfection with mitochondrial targeted catalase inhibited mitochondrial ROS production and potentiated cell death. These data show that ROS production is important in receptor-mediated apoptosis and may play a pivotal role in cell survival.     

TCDD decreases ATP levels and increases reactive oxygen production through changes in mitochondrial F(0)F(1)-ATP synthase and ubiquinone

Mitochondria generate ATP and participate in signal transduction and cellular pathology and/or cell death. TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) decreases hepatic ATP levels and generates mitochondrial oxidative DNA damage, which is exacerbated by increasing mitochondrial glutathione redox state and by inner membrane hyperpolarization. This study identifies mitochondrial targets of TCDD that initiate and sustain reactive oxygen production and decreased ATP levels. One week after treating mice with TCDD, liver ubiquinone (Q) levels were significantly decreased, while rates of succinoxidase and Q-cytochrome c oxidoreductase activities were increased. However, the expected increase in Q reduction state following TCDD treatment did not occur; instead, Q was more oxidized. These results could be explained by an ATP synthase defect, a premise supported by the unusual finding that TCDD lowers ATP/O ratios without concomitant changes in respiratory control ratios. Such results suggest either a futile cycle in ATP synthesis, or hydrolysis of newly synthesized ATP prior to release. The TCDD-mediated decrease in Q, concomitant with an increase in respiration, increases complex 3 redox cycling. This acts in concert with glutathione to increase membrane potential and reactive oxygen production. The proposed defect in ATP synthase explains both the greater respiratory rates and the lower tissue ATP levels.