RESUMO
Non-protein bound iron (NPBI) is able to catalyse oxidative reactions, causing damage to vital structures. Adverse effects induced by cisplatin seem, in part, to be mediated by free radicals. In the present study, we have measured plasma NPBI, various other iron parameters and antioxidants in 28 cancer patients undergoing cisplatin-based chemotherapy at various time points before and during chemotherapy. No NPBI was present prior to therapy, but within 1-4 days following the first administration of chemotherapy, mean NPBI rose significantly to 10.6+/-6.6 micromol/l (range, 0.6-21.3 micromol/l) in 18 (64.3%) of the 28 patients measured. The rise in NPBI was accompanied by a significant rise in total plasma iron and ferritin and a marked decrease in the latent iron-binding capacity. Concomitantly, plasma vitamins C and E decreased significantly, indicating consumption of antioxidants. Similar observations were also made during the fourth chemotherapy cycle. The increase in NPBI preceded and correlated significantly with chemotherapy toxicity, such as a decrease in leucocyte count and haemoglobin, with a transient rise in various liver enzymes and with known cisplatin-related toxicity, i.e. the loss of renal and hearing function. In conclusion, cisplatin chemotherapy induces oxidative damage which rapidly leads to release of iron from intracellular proteins and the appearance of NPBI. Bone marrow, red blood cells, liver and kidney seem to be a likely source of NPBI. The observed high levels of NPBI may be a major causative determinant in chemotherapy-induced toxicity.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Germinoma/tratamento farmacológico , Ferro/sangue , Adolescente , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Antioxidantes/metabolismo , Ácido Ascórbico/sangue , Ácido Ascórbico/uso terapêutico , Cisplatino/administração & dosagem , Cisplatino/efeitos adversos , Feminino , Ferritinas/sangue , Germinoma/sangue , Audição/efeitos dos fármacos , Humanos , Proteínas de Ligação ao Ferro , Rim/efeitos dos fármacos , Rim/fisiopatologia , Fígado/efeitos dos fármacos , Fígado/fisiopatologia , Masculino , Malondialdeído/sangue , Pessoa de Meia-Idade , Vitamina E/sangue , Vitamina E/uso terapêuticoRESUMO
PURPOSE: To study the protective effect of topically applied vitamin E (TOC), vitamin C (ASC), or a combination of both, against the lipid peroxidation of eicosapentaenoic acid (EPA) induced by simulated solar radiation (SSR). MATERIAL AND METHODS: EPA (25 nmol cm(-2)) was topically applied to pig skin explants, followed by increasing doses of TOC and ASC, either alone or combined. Epidermal lipid peroxidation was assessed after 15 min of exposure to SSR (resulting in a UVA and UVB dose of 18 and 3 kJ m(-2), respectively). SSR-induced changes in the levels of TOC and ASC were determined in the stratum cornaeum and the viable epidermis. RESULTS: SSR exposure of EPA-treated pig skin resulted in a twofold increase in epidermal lipid peroxidation (p <0.005) which was reduced by topically applied TOC or ASC 60 min before SSR exposure (p <0.05). Compared with TOC (5 nmol cm(-2)), a 400-fold higher ASC dose was needed and only TOC provided complete protection against the lipid peroxidation of EPA. The levels of both TOC and ASC clearly increased in both skin compartments by increasing the applied dose of these two compounds (p <0.05). In contrast to ASC, TOC was consumed by up to 55-70% during SSR exposure (p <0.05). Compared with separate application, combined TOC and ASC efficiently protected against lipid peroxidation of EPA at doses that were five and 200 times lower, respectively. In the presence of low ASC doses, 70-100% of epidermal TOC was regenerated during SSR exposure (p <0.05). CONCLUSIONS: Topically applied TOC and ASC protect against SSR-induced lipid peroxidation of EPA. The synergism between TOC and ASC resulted in a more efficient protection at substantially lower doses of both antioxidants. Co-supplementation of EPA with TOC and/or ASC might improve the beneficial biological effects of EPA.
Assuntos
Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Ácido Eicosapentaenoico/metabolismo , Pele/efeitos dos fármacos , Pele/efeitos da radiação , Vitamina E/farmacologia , Administração Tópica , Animais , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Peroxidação de Lipídeos , Energia Solar , Suínos , Fatores de Tempo , Raios UltravioletaRESUMO
Moison, R. M. W. and Beijersbergen van Henegouwen, G. M. J. Dietary Eicosapentaenoic Acid Prevents Systemic Immunosuppression in Mice Induced by UVB Radiation. Radiat. Res. 156, 36-44 (2001). Reactive oxygen species (ROS) contribute to the immunosuppression induced by UVB radiation. Omega-3 fatty acids in fish oil, e.g. eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), can modulate immunoresponsiveness, but because of their susceptibility to ROS-induced damage, they can also challenge the epidermal antioxidant defense system. The influence of dietary supplementation with different omega-3 fatty acids on systemic immunosuppression induced in mice by UVB radiation was studied using the contact hypersensitivity response to trinitrochlorobenzene. In an attempt to study the mechanisms involved, UVB-radiation-induced changes in epidermal antioxidant status were also studied. Mice received high-fat (25% w/w) diets enriched with either oleic acid (control diet), EPA, DHA, or EPA + DHA (MaxEPA). Immunosuppression induced by UVB radiation was 53% in mice fed the oleic acid diet and 69% in mice fed the DHA diet. In contrast, immunosuppression was only 4% and 24% in mice fed the EPA and MaxEPA diets, respectively. Increased lipid peroxidation and decreased vitamin E levels (P < 0.05) were found in unirradiated mice fed the MaxEPA and DHA diets. For all diets, exposure to UVB radiation increased lipid peroxidation (P < 0.05), but levels of glutathione (P < 0.05) and vitamin C (P > 0.05) decreased only in the mice given fish oil. UVB irradiation did not influence vitamin E levels. In conclusion, dietary EPA, but not DHA, protects against UVB-radiation-induced immunosuppression in mice. The degree of protection appears to be related to the amount of EPA incorporated and the ability of the epidermis to maintain an adequate antioxidant level after irradiation.
Assuntos
Ácido Eicosapentaenoico/farmacologia , Epiderme/efeitos da radiação , Alimentos Formulados , Tolerância Imunológica/efeitos dos fármacos , Tolerância Imunológica/efeitos da radiação , Raios Ultravioleta/efeitos adversos , Animais , Antioxidantes/metabolismo , Antioxidantes/efeitos da radiação , Ácido Ascórbico/metabolismo , Epiderme/efeitos dos fármacos , Epiderme/imunologia , Epiderme/metabolismo , Ácidos Graxos/análise , Alimentos Formulados/análise , Glutationa/metabolismo , Tolerância Imunológica/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Cloreto de Picrila/imunologia , Cloreto de Picrila/farmacologia , Vitamina E/metabolismoRESUMO
BACKGROUND: Prooxidant activity of non-protein-bound iron (NPBI) is an important contributor to reactive oxygen species-induced injury after the resuscitation of critically ill patients. Plasma NPBI occurs in critically ill adults, children, and newborn babies, who often require resuscitation. The ability of the resuscitation fluids to bind iron and lower the patients' NPBI levels in vitro has not previously been studied. STUDY DESIGN AND METHODS: In an in vitro model, highly iron-saturated cord blood plasma from 10 preterm and 10 term babies was mixed with FFP, pasteurized plasma protein solution, and 0.9-percent saline. Plasma from 10 healthy adult volunteers was used as a control. Before and after the mixing with any resuscitation fluid, NPBI levels and ceruloplasmin iron-oxidizing and transferrin iron-binding antioxidant capacities were measured. RESULTS: After the in vitro mixing with FFP, the incidence and concentration of NPBI were markedly decreased and the iron-binding antioxidant capacity was increased in the plasma of the preterm and term babies. Being mixed with pasteurized plasma protein solution and 0.9-percent saline did not influence the iron-binding antioxidant capacity of newborn babies' plasma. In the control plasma, results were not changed after the mixing with any resuscitation fluid. In every group, the iron-oxidizing antioxidant capacity was not changed after the mixing with any fluid. CONCLUSION: Iron-induced oxidative tissue damage may be influenced by resuscitation fluids. In the ongoing debate over the choice of crystalloid or colloid resuscitation fluids, the influence of each fluid on the patient's antioxidant capacity warrants more attention.
Assuntos
Antioxidantes/farmacologia , Transfusão de Sangue , Ferro/sangue , Ressuscitação , Adulto , Feminino , Humanos , Recém-Nascido , Recém-Nascido Prematuro/sangue , Ferro/metabolismo , Oxirredução , Gravidez , Ligação ProteicaRESUMO
To analyze the effects of radiochemotherapy on the pro-oxidative/antioxidative balance in plasma, we measured the total radical antioxidant parameter of plasma (TRAP) and single plasma antioxidants (uric acid, sulfhydryl groups, alpha-tocopherol, ubiquinone-10/total coenzyme-Q10 ratio, ascorbate, and bilirubin) every 12 h during high-dose chemotherapy and radiochemotherapy preceding bone marrow transplantation (BMT). Nontransferrin-bound iron (NTBI) was monitored as a potential pro-oxidant. Plasma levels of polyunsaturated fatty acids (PUFA) were measured as substrates, and thiobarbituric acid-reactive substances (TBARS) were measured as products of lipid peroxidation. Allantoin was analyzed as the product of uric acid oxidation. Patients receiving busulfan, VP-16, and cyclophosphamide (BU/VP/CY) (n = 8) were compared with those receiving total body irradiation in addition to VP-16 and cyclophosphamide (TBI/VP/CY) (n = 8). TRAP values were within the normal range before therapy and decreased after BU/VP/CY by 37% (p <. 02) and after TBI/VP/CY by 39% (p <.02). During TBI and after VP-16, a temporary increase in TRAP values occurred, which was not related to changes in individual antioxidants. In vitro experiments confirmed that VP-16 had an antioxidative effect. The concentration of uric acid declined in both groups and correlated with TRAP (BU/VP/CY: r =.80, p <.001; TBI/VP/CY: r =.84, p <.001). Levels of NTBI, which is normally not found in plasma, increased rapidly during conditioning therapy (p <.02 in both groups) and correlated inversely with TRAP (weighted intraindividual Spearman rank correlation coefficient for both groups: NTBI and TRAP: r = -.59, p <.001) and PUFA (in the radiochemotherapy group: r = -.67, p <.001). Whereas PUFA declined (p <.02 in both groups), TBARS increased (p <. 05 in both groups). Furthermore, an increase of allantoin and ubiquinone-10/total coenzyme-Q10 ratio in the BU/VP/CY group was found (allantoin: p <.02; ubiquinone-10/total coenzyme-Q10 ratio: p <.05). Antioxidants only partially recovered to baseline values until day 14 after BMT. Our findings indicate oxidative stress after high-dose radiochemotherapy and suggest a contribution of NTBI therein.
Assuntos
Antioxidantes/metabolismo , Transplante de Medula Óssea , Neoplasias Hematológicas/terapia , Ferro/sangue , Alantoína/sangue , Antineoplásicos/uso terapêutico , Bussulfano/uso terapêutico , Terapia Combinada , Ciclofosfamida/uso terapêutico , Etoposídeo/uso terapêutico , Ácidos Graxos Insaturados/sangue , Neoplasias Hematológicas/sangue , Neoplasias Hematológicas/radioterapia , Humanos , Peróxidos Lipídicos/sangue , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Estatísticas não Paramétricas , Ubiquinona/sangue , Ácido Úrico/sangue , Irradiação Corporal TotalRESUMO
OBJECTIVE: Free radical-induced postasphyxial reperfusion injury has been recognized as an important cause of brain tissue damage. We investigated the effect of high-dose allopurinol (ALLO; 40 mg/kg), a xanthine-oxidase inhibitor and free radical scavenger, on free radical status in severely asphyxiated newborns and on postasphyxial cerebral perfusion and electrical brain activity. METHODS: Free radical status was assessed by serial plasma determination of nonprotein-bound iron (microM), antioxidative capacity, and malondialdehyde (MDA; microM). Cerebral perfusion was investigated by monitoring changes in cerebral blood volume (delta CBV; mL/100 g brain tissue) with near infrared spectroscopy; electrocortical brain activity (ECBA) was assessed in microvolts by cerebral function monitor. Eleven infants received 40 mg/kg ALLO intravenously, and 11 infants served as controls (CONT). Plasma nonprotein-bound iron, antioxidative capacity, and MDA were measured before 4 hours, between 16 and 20 hours, and at the second and third days of age. Changes in CBV and ECBA were monitored between 4 and 8, 16 and 20, 58 and 62, and 104 and 110 hours of age. RESULTS: Six CONT and two ALLO infants died after neurologic deterioration. No toxic side effects of ALLO were detected. Nonprotein-bound iron (mean +/- SEM) in the CONT group showed an initial rise (18.7 +/- 4.6 microM to 21.3 +/- 3.4 microM) but dropped to 7.4 +/- 3.5 microM at day 3; in the ALLO group it dropped from 15.5 +/- 4.6 microM to 0 microM at day 3. Uric acid was significantly lower in ALLO-treated infants from 16 hours of life on. MDA remained stable in the ALLO group, but increased in the CONT group at 8 to 16 hours versus < 4 hours (mean +/- SEM; 0.83 +/- 0.31 microM vs 0.50 +/- 0.14 microM). During 4 to 8 hours, delta CBV-CONT showed a larger drop than delta CBV-ALLO from baseline. During the subsequent registrations CBV remained stable in both groups. ECBA-CONT decreased, but ECBA-ALLO remained stable during 4 to 8 hours of age. Neonates who died had the largest drops in CBV and ECBA. CONCLUSION: This study suggests a beneficial effect of ALLO treatment on free radical formation, CBV, and electrical brain activity, without toxic side effects.
Assuntos
Alopurinol/uso terapêutico , Antimetabólitos/uso terapêutico , Asfixia Neonatal/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Circulação Cerebrovascular/efeitos dos fármacos , Radicais Livres/metabolismo , Alopurinol/sangue , Alopurinol/farmacologia , Antimetabólitos/sangue , Antimetabólitos/farmacologia , Asfixia Neonatal/metabolismo , Asfixia Neonatal/fisiopatologia , Encéfalo/irrigação sanguínea , Encéfalo/fisiopatologia , Eletroencefalografia/efeitos dos fármacos , Eletrofisiologia , Hemodinâmica/efeitos dos fármacos , Humanos , Recém-Nascido , Peroxidação de Lipídeos/efeitos dos fármacosRESUMO
We compared the postnatal changes (days 1-28) in red blood cell glutathione and plasma sulfhydryl content in preterm babies developing chronic lung disease (CLD, n = 13) to those in babies with respiratory distress syndrome (RDS, n = 13) and control babies (n = 21). There were no initial differences in these measurements between the three groups. However, on day 28 in CLD and RDS the red blood cell glutathione was decreased compared to the control babies (p < 0.05). In CLD, there was a significant correlation between reduced/oxidized glutathione and (i) maximal FiO2 (r = -0.69, p < 0.05) and (ii) minimal a/A ratio (r = +0.73, p < 0.005). On day 28, although the plasma sulfhydryl level did not differ between the groups, the sulfhydryl/total protein ratio was decreased in CLD (p < 0.05). The late decrease in both glutathione and sulfhydryl/total protein ratio in babies with CLD suggests that oxidative stress is still ongoing at 28 days after birth and that the antioxidant capacity of their blood is still diminished at this time.
Assuntos
Displasia Broncopulmonar/sangue , Eritrócitos/metabolismo , Glutationa/sangue , Recém-Nascido Prematuro/sangue , Síndrome do Desconforto Respiratório do Recém-Nascido/sangue , Compostos de Sulfidrila/sangue , Análise de Variância , Displasia Broncopulmonar/diagnóstico , Displasia Broncopulmonar/terapia , Feminino , Glutationa/metabolismo , Humanos , Recém-Nascido , Masculino , Oxigênio/uso terapêutico , Plasma/metabolismo , Valores de Referência , Respiração Artificial , Síndrome do Desconforto Respiratório do Recém-Nascido/diagnóstico , Síndrome do Desconforto Respiratório do Recém-Nascido/terapia , Compostos de Sulfidrila/metabolismoRESUMO
We investigated in an in vitro transfusion model the early effects of plasma preparations and donor red blood cells on the antioxidant capacity of the cord blood from babies. Addition of pasteurized plasma protein solution to plasma from babies decreased the peroxyl radical trapping capacity (p < 0.02). In contrast, fresh frozen plasma did not lower this capacity. Addition of adult donor red blood cells to the babies' red blood cells did not significantly decrease the glutathione-recycling capacity of the blood. On the basis of these in vitro results we hypothesize that the use of resuscitation fluids with low antioxidant capacity may temporarily decrease the ability of the baby to catabolize reactive oxygen species.
Assuntos
Doadores de Sangue , Transfusão de Eritrócitos , Recém-Nascido , Plasma/fisiologia , Adulto , Técnicas de Cultura de Células , Sangue Fetal , Glutationa/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Plasma/efeitos dos fármacos , EspectrofotometriaRESUMO
Preformed lipid peroxidation products present in the feed may contribute to the total reactive oxygen radical load infants have to deal with and may play a role in the pathogenesis of necrotizing enterocolitis and bronchopulmonary dysplasia. In this study, the occurrence of lipid peroxidation in human milk and feeding formulas for preterm babies was evaluated in vitro. Free linoleic acid (18:2) and its hydroperoxide (18:2OOH) were measured by gas chromatography-mass spectrometry and the concentration of 18:2OOH and the 18:2OOH/18:2 ratio were used as indices of peroxidation. In all feeds peroxidation products were present, but the proportion of peroxidized 18:2 was greater in infant formula. Storage of human milk (+4 degrees C for four days) increased lipid peroxidation. Exposure to light during tube feeding increased peroxidation in infant formula but not in human milk. Different procedures for preparation, storage and feeding may decrease the concentration of these potentially toxic peroxidized lipids in human milk and infant formula.
Assuntos
Nutrição Enteral , Manipulação de Alimentos , Alimentos Infantis/efeitos da radiação , Peroxidação de Lipídeos/efeitos da radiação , Leite Humano/metabolismo , Leite Humano/efeitos da radiação , Fototerapia , Cromatografia Gasosa-Espectrometria de Massas/instrumentação , Cromatografia Gasosa-Espectrometria de Massas/métodos , Humanos , Lactente , Alimentos Infantis/análise , Ácido Linoleico , Ácidos Linoleicos/análise , Ácidos Linoleicos/efeitos da radiação , Leite Humano/químicaRESUMO
The purpose of this study was to examine the role of glutathione depletion and alterations in the energy status in the induction of acute cytotoxicity to freshly isolated rat hepatocytes. Depletion of intracellular glutathione by diethyl maleate and phorone to levels below 5% of control did not induce loss of viability nor loss of intracellular ATP. Ethacrynic acid, a compound known to deplete mitochondrial GSH in addition to cytosolic GSH, induced cell killing after a depletion of ATP, next to GSH depletion. The results confirmed that depletion of intracellular glutathione alone does not necessarily result in cell killing. Only when glutathione depletion is succeeded by reduction in ATP levels, loss of cell viability is observed. The relationship between alterations in the energy status and the induction of cell death was further substantiated by inhibition of glycolytic and mitochondrial ATP generation. Treatment of hepatocytes either with iodoacetic acid to inhibit glycolysis (in hepatocytes from fed rats) or with potassium cyanide to inhibit mitochondrial respiration (in hepatocytes from both fed and fasted rats) revealed that depletion of intracellular ATP could lead to lethal cell injury. The susceptibility of cells to metabolic inhibition was better reflected by the rate of reduction in the energy charge than by the reduction of ATP alone. In conclusion, our results suggest that alterations of the energy status may be a critical event in the induction of irreversible cell injury. Depletion of cellular GSH is only cytotoxic when followed by a reduction of the energy charge.
Assuntos
Trifosfato de Adenosina/metabolismo , Glutationa/metabolismo , Fígado/citologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ácido Etacrínico/farmacologia , Cetonas/farmacologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Maleatos/farmacologia , Cianeto de Potássio/farmacologia , Ratos , Ratos WistarRESUMO
The ability of erythrocytes from newborn babies and adults to maintain reduced glutathione levels during oxidative stress was studied. In vitro incubation of erythrocytes with H2O2, with or without inactivation of catalase, caused a rapid depletion of reduced glutathione (GSH) and concomitant accumulation of oxidized glutathione followed by recovery of GSH and fall of oxidized glutathione to initial values in all subjects. Inactivation of catalase resulted in a 50% loss of intracellular glutathione (p less than 0.005), a larger maximum GSH depletion (p less than 0.05), and a longer GSH recovery time (p less than 0.005). Erythrocytes from newborn babies showed a smaller maximum GSH depletion (p less than 0.05) and a shorter GSH recovery time (p less than 0.005) compared with those from adults. These differences between the newborn and adult groups persisted after inactivation of catalase. An increase in maximum GSH depletion and GSH recovery time (p less than 0.005) was observed when a lower hematocrit was used for these GSH recovery studies. Effective glutathione recycling in erythrocytes may protect immature tissues of the newborn baby from peroxidative damage.
Assuntos
Eritrócitos/metabolismo , Glutationa/sangue , Recém-Nascido/sangue , Adulto , Catalase/sangue , Eritrócitos/efeitos dos fármacos , Glutationa/análogos & derivados , Dissulfeto de Glutationa , Humanos , Peróxido de Hidrogênio/farmacologia , Técnicas In Vitro , Oxirredução , Espécies Reativas de Oxigênio/metabolismo , Estresse Fisiológico/sangueRESUMO
In this study the effect of metabolism of menadione (2-methyl-1,4-naphthoquinone) on ATP generation in isolated rat hepatocytes was investigated. Menadione-induced cytotoxicity correlated well with the depletion of ATP. Loss of viability lagged approximately 25 min behind the depletion of ATP. Our results suggest that depletion of ATP may be mediated by interference with glycolysis and protein breakdown, resulting in a lack of oxidizable substrates for ATP generation. (i) Menadione reduced proteolysis to 27% of control after 60 min of incubation. (ii) Increased glycogenolysis was not accompanied by accumulation of glycolytic end-products. The increased levels of glucose 6-phosphate were mainly metabolized to glucose. (iii) Menadione induced a time- and concentration-dependent inhibition of the glyceraldehyde-3-phosphate dehydrogenase activity, although no accumulation of glycolytic intermediates was found. The data presented suggest that glycolysis may be inhibited upstream of glyceraldehyde-3-phosphate dehydrogenase. (iv) Suppletion of metabolic substrates (pyruvate, oxaloacetate, and glutamine) postponed the menadione-induced ATP depletion and delayed the onset of cell killing. The protecting effect of these metabolic substrates could be reversed by atractyloside, an inhibitor of the ADP/ATP translocase. The temporary protection of metabolic substrates suggests that additional mechanisms (e.g., cofactor depletion, mitochondrial damage, enzyme inactivation) may play a role in menadione-induced ATP depletion. The present study substantiates the critical role of ATP depletion in menadione-induced cell death.
Assuntos
Trifosfato de Adenosina/metabolismo , Fígado/metabolismo , Vitamina K/farmacologia , Animais , Atractilosídeo/farmacologia , Células Cultivadas , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Glicólise/efeitos dos fármacos , Cinética , Fígado/efeitos dos fármacos , Glicogênio Hepático/metabolismo , Masculino , Proteínas/metabolismo , Ratos , Ratos Endogâmicos , Vitamina K/metabolismoRESUMO
The biochemical mechanism of cytotoxicity, induced by the quinoid compound 2-methyl 1,4-naphthoquinone (menadione), was investigated in hepatocytes freshly isolated from fasted and fed rats. Hepatocytes from fasted rats were significantly more vulnerable to the toxicity of menadione than hepatocytes from fed rats. Menadione (150 microM) induced a 50% loss of viability of cells (LT50) from fasted rats after 55 min of incubation, whereas a LT50 of 80 min was observed after exposure of hepatocytes from fed rats to menadione. Glutathione and NADPH levels were rapidly depleted by menadione metabolism. This depletion was sustained during the incubation period. No significant differences were found in the time course and extent of the menadione-induced glutathione and NADPH depletion in hepatocytes of both nutritional states. Menadione also affected the energy status of the hepatocytes. The ATP content of cells from fasted rats decreased to 50% (AT50) within 18 min of exposure to menadione, whereas a 50% loss of ATP content of hepatocytes from fed rats was reached at 65 min. In contrast to depletion of glutathione and NADPH, the time course and extent of menadione-induced ATP depletion correlated well with the time of onset and rate of cell killing. Our results suggest that menadione metabolism may interfere with both mitochondrial and glycolytic ATP production. Depletion of ATP might be a critical step in menadione-induced cytotoxicity.