Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P42574 (caspase-3)
 45,978 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have recently shown that dithiocarbamate (DC) disulfides inhibit proteolytic processing of the caspase-3 proenzyme in Jurkat T lymphocytes treated with anti-CD95 (Fas/APO-1) antibody. Because the processing can be accomplished by caspase activity, we investigated the effect of DC disulfides, such as disulfiram (DSF), on active caspases. DSF showed a dose-dependent inhibition was prevented by including dithiothreitol (DTT) in the reaction buffer, thiol-disulfide exchange between inhibitor and target is suggested. Direct interaction of DSF with caspases was confirmed by its inhibition of the purified Ac-DEVD-AMC cleaving protease, caspase-3 (CPP32/apopain). An apparent rate constant (K(app)) for this inhibition was estimated to be 0.45 x 10(3)M(-1)s(-1). DSF was also observed to inhibit the purified Ac-YVAD-AMC cleaving enzyme, caspase-1 (interleukin-1 beta-converting enzyme, ICE), with a K(app) of 2.2 x 10(3) M(-1)s(-1). In this case protein mixed disulfide formation between DSF and caspase-1 was directly demonstrated using 35S-labeled DSF. The physiological disulfide GSSG was also observed to influence the activity of caspases. A glutathione buffer (5 mM) with a GSH:GSSG ratio of 9:1 decreased the Ac-DEVD-AMC cleaving activity in S100 cytosolic extracts by 50% as compared to GSH controls without GSSG. In conclusion, our study shows that caspases are quite sensitive to thiol oxidation and that DSF is a very potent oxidant of caspase protein thiol(s), being 700-fold more potent than glutathione disulfide.
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PMID:Disulfiram is a potent inhibitor of proteases of the caspase family. 943 20

The purpose of this study was to determine if exacerbation of apoptosis precedes liver injury during chronic exposure of rats to alcohol. After 7 weeks of feeding an alcohol- or dextrin-containing liquid diet, the animals were treated with gram-negative bacterial lipopolysaccharide (1 mg x kg(-1) body weight, intravenously) or sterile saline and sacrificed 3 hr after the treatment. Alanine:2-oxoglutarate aminotransferase (ALT) and lactate:NAD oxidoreductase [lactate dehydrogenase (LDH)] were measured in plasma. The caudate lobe of the liver was resected for histology, while the rest of the organ was perfused with collagenase to isolate hepatocytes, Kupffer cells (KCs), and sinusoidal endothelial cells (SECs) by centrifugal elutriation. Hepatocyte mitochondria were isolated by differential centrifugation of the cell homogenate. Reduced and oxidized glutathione (GSH and GSSG) in isolated hepatocytes and hepatocyte mitochondria, and malondialdehyde in hepatocytes were assayed. Caspase-3 activity and Fas ligand mRNA expression were determined in hepatocytes, KCs, and SECs. Plasma ALT and LDH activity, liver histology, GSH, GSSG and their ratio, and malondialdehyde content were not affected by alcohol treatment Caspase-3 activity was significantly increased in alcohol-treated rats in all three cell types, with the lowest response observed in hepatocytes and the highest in KCs. Fas ligand mRNA expression, which had the highest level in SECs, followed by KCs and hepatocytes, was not affected by alcohol administration. Lipopolysaccharide had the following effects: an increase in ALT in both pair- and alcohol-fed rats, and LDH only in alcohol-fed rats, a decrease in GSH + GSSG levels in both mitochondria and hepatocytes, an elevation of malondialdehyde content in hepatocytes, a raise in caspase-3 activity in all groups and cell types, and an augmentation of Fas ligand expression in hepatocytes and KCs, but not in SECs. These data suggest that, during chronic alcohol consumption, an exacerbated apoptosis precedes alcohol-induced liver injury.
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PMID:Modulation of caspase-3 activity and Fas ligand mRNA expression in rat liver cells in vivo by alcohol and lipopolysaccharide. 1006 67

Agonistic engagement of the cytokine receptor CD95 in mice leads to activation of hepatic caspases, followed by massive hepatocyte apoptosis, acute liver failure, and death. This mechanism of cell death is thought to be associated with several human liver disorders. Because hepatic glutathione represents the major defense against toxic liver injury, we investigated its role in CD95-mediated liver failure, which represents a model for hyperinflammatory organ destruction. As a tool for modulating the liver glutathione status of mice in vivo, we used the GSH transferase substrate, phorone, which rapidly depleted hepatic glutathione in a dose-dependent manner. When GSH was depleted, CD95-initiated hepatic caspase-3-like activity and DNA fragmentation were completely blocked, and animals were protected from liver injury dose-dependently as assessed by histological examination and determination of liver enzymes in plasma. Conversely, repletion of hepatic glutathione by treatment with the permeable glutathione monoethylester restored susceptibility of GSH-depleted mice toward CD95-mediated liver injury. In contrast, the antioxidants, GSH, N-acetyl cysteine, alpha-tocopherol, butyl-hydroxytoluene, and catalase failed to do so. Animals treated once with phorone survived for more than 3 months after an otherwise lethal injection of the activating anti-CD95 antibody. We investigated the thiol sensitivity of recombinant caspase-3 in vitro and observed that its activity was dependent on the presence of a reducing agent such as GSH, while GSSG attenuated proteolytic activity. Based on our finding that CD95-mediated hepatocyte apoptosis requires an intact intracellular glutathione status, we propose that the activation of apoptosis-executing caspases is controlled by reduced glutathione.
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PMID:CD95-Mediated murine hepatic apoptosis requires an intact glutathione status. 1038 54

Mitochondria serve as a pivotal component of the apoptotic cell death machinery. However, cells that lack mitochondrial DNA (rho(0) cells) retain apparently normal apoptotic signaling. In the present study, we examined mitochondrial mechanisms of apoptosis in rho(0) osteosarcoma cells treated with staurosporine. Immunohistochemistry revealed that rho(0) cells maintained a normal cytochrome c distribution in mitochondria even though these cells were deficient in respiration. Upon staurosporine treatment, cytochrome c was released concomitantly with activation of caspase 3 and loss of mitochondrial membrane potential (Deltapsi(m)). After mitochondrial loss of cytochrome c, rho(0) cells underwent little change in glutathione (GSH) redox potential whereas a dramatic oxidation in GSH/glutathione disulfide (GSSG) pool occurred in parental rho(+) cells. These results show that mitochondrial signaling of apoptosis via cytochrome c release was preserved in cells lacking mtDNA. However, intracellular oxidation that normally accompanies apoptosis was lost, indicating that the mitochondrial respiratory chain provides the major source of redox signaling in apoptosis.
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PMID:Cytochrome c-mediated apoptosis in cells lacking mitochondrial DNA. Signaling pathway involving release and caspase 3 activation is conserved. 1051 72

Recent results demonstrated that S-nitrosoglutathione (GSNO) and nitric oxide (*NO) protect brain dopamine neurons from hydroxyl radical (*OH)-induced oxidative stress in vivo because they are potent antioxidants. GSNO and *NO terminate oxidant stress in the brain by (i) inhibiting iron-stimulated hydroxyl radicals formation or the Fenton reaction, (ii) terminating lipid peroxidation, (iii) augmenting the antioxidative potency of glutathione (GSH), (iv) mediating neuroprotective action of brain-derived neurotrophin (BDNF), and (v) inhibiting cysteinyl proteases. In fact, GSNO--S-nitrosylated GSH--is approximately 100 times more potent than the classical antioxidant GSH. In addition, S-nitrosylation of cysteine residues by GSNO inactivates caspase-3 and HIV-1 protease, and prevents apoptosis and neurotoxicity. GSNO-induced antiplatelet aggregation is also mediated by S-nitrosylation of clotting factor XIII. Thus the elucidation of chemical reactions involved in this GSNO pathway (GSH GS* + *NO-->[GSNO]-->GSSG + *NO-->GSH) is necessary for understanding the biology of *NO, especially its beneficial antioxidative and neuroprotective effects in the CNS. GSNO is most likely generated in the endothelial and astroglial cells during oxidative stress because these cells contain mM GSH and nitric oxide synthase. Furthermore, the transfer of GSH and *NO to neurons via this GSNO pathway may facilitate cell to neuron communications, including not only the activation of guanylyl cyclase, but also the nitrosylation of iron complexes, iron containing enzymes, and cysteinyl proteases. GSNO annihilates free radicals and promotes neuroprotection via its c-GMP-independent nitrosylation actions. This putative pathway of GSNO/GSH/*NO may provide new molecular insights for the redox cycling of GSH and GSSG in the CNS.
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PMID:The redox pathway of S-nitrosoglutathione, glutathione and nitric oxide in cell to neuron communications. 1063 Jun 87

4-Hydroxynonenal (HNE), a diffusible product of lipid peroxidation, has been suggested to be a key mediator of oxidative stress-induced cell death. In this study, we partially characterized the mechanism of HNE-mediated cytotoxicity. Incubation of human T lymphoma Jurkat cells with 20-50 microM HNE led to cell death accompanied by DNA fragmentation. Western blot analysis showed that HNE-treatment induced time- and dose-dependent activation of caspase-8, caspase-9 and caspase-3. HNE-induced caspase-3 processing was confirmed by a flow cytometric demonstration of increased catalytic activity on the substrate peptide. HNE treatment also led to remarkable cleavage of poly(ADP-ribose) polymerase (PARP), which was prevented by pretreatment of cells with DEVD-FMK as a caspase-3 inhibitor. The HNE-mediated activation of caspases, cleavage of PARP and DNA fragmentation were blocked by antioxidants cysteine, N-acety-L-cysteine and dithiothreitol, but not by two other HNE-reactive amino acids lysine and histidine, or by cystine, the oxidized form of cysteine. HNE rapidly decreased levels of intracellular reduced glutathione (GSH) and its oxidized form GSSG, and these were also attenuated by the reductants. Coincubation of Jurkat cells with a blocking anti-Fas antibody prevented Fas-induced but not HNE-induced activation of caspase-3. HNE also activated caspase-3 in K562 cells that do not express functional Fas. Our results thereby demonstrate that HNE triggers oxidative stress-linked apoptotic cell death through activation of the caspase cascade. The results also suggest a possible mechanism involving a direct scavenge of intracellular GSH by HNE.
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PMID:4-hydroxynonenal induces a cellular redox status-related activation of the caspase cascade for apoptotic cell death. 1065 56

Both caspase-3 and -6-like activities increased in the cytosolic extract from ricin-treated U937 cells that were inhibited by glutathione disulfide (GSSG) in a dose-dependent manner, but reduced glutathione (GSH) had no effect. Interestingly, caspase-6 like activity was more sensitive to GSSG than caspase-3 like activity. The IC50 of GSSG against caspase-3 and caspase-6 like activities were estimated to be 2.8 mM and 0.8 mM, respectively. Cystine but not cysteine also showed similar inhibitory effect on caspase-3-like activity. The inhibitory effect of GSSG on these caspase-like activities was prevented by the addition of DTT to the assay mixture. These results suggest that an intact disulfide portion of GSSG is required for the effective inhibition of caspase activity.
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PMID:Effects of glutathione-related compounds on increased caspase-3 and caspase-6-like activities in ricin-treated U937 cells. 1076 9

Apoptosis plays a critical role in maintaining homeostasis of the intestinal epithelium. Dietary oxidants like peroxidized lipids could perturb cellular redox status and disrupt mucosal turnover. The objective of this study was to delineate the role of lipid hydroperoxide (LOOH) -induced redox shifts in intestinal apoptosis using the human colonic CaCo-2 cell. We found that subtoxic concentrations of LOOH increased CaCo-2 cell apoptosis. This LOOH-induced apoptosis was associated with a significant decrease in the ratio of reduced glutathione-to-oxidized glutathione (GSH/GSSG), which preceded DNA fragmentation by 12 to 14 h, suggesting a temporal relationship between the two events. Oxidation of GSH with the thiol oxidant diamide caused significant decreases in cellular GSH and GSH/GSSG at 15 min that correlated with the activation of caspase 3 (60 min) and cleavage of PARP (120 min), confirming a temporal link between induction of cellular redox imbalance and initiation of apoptotic cell death. These kinetic studies further reveal that oxidant-mediated early redox change (within 1 h) was a primary inciting event of the apoptotic cascade. Once initiated, the recovery of redox balance did not prevent the progression of CaCo-2 cell apoptosis to its biological end point at 24 h. Collectively, the study shows that subtoxic levels of LOOH disrupt intestinal redox homeostasis, which contributes to apoptosis. These results provide insights into the mechanism of hydroperoxide-induced mucosal turnover that have important implications for understanding oxidant-mediated genesis of gut pathology.
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PMID:Lipid hydroperoxide-induced apoptosis in human colonic CaCo-2 cells is associated with an early loss of cellular redox balance. 1092 91

Oxidative injuries including apoptosis can be induced by reactive oxygen species (ROS) and reactive nitrogen species (RNS) in aerobic metabolism. We determined impacts of a selenium-dependent glutathione peroxidase-1 (GPX1) on apoptosis induced by diquat (DQ), a ROS (superoxide) generator, and peroxynitrite (PN), a potent RNS. Hepatocytes were isolated from GPX1 knockout (GPX1-/-) or wild-type (WT) mice, and treated with 0.5 mm DQ or 0.1-0.8 mm PN for up to 12 h. Loss of cell viability, high levels of apoptotic cells, and severe DNA fragmentation were produced by DQ in only GPX1-/- cells and by PN in only WT cells. These two groups of cells shared similar cytochrome c release, caspase-3 activation, and p21(WAF1/CIP1) cleavage. Higher levels of protein nitration were induced by PN in WT than GPX1-/- cells. Much less and/or slower cellular GSH depletion was caused by DQ or PN in GPX1-/- than in WT cells, and corresponding GSSG accumulation occurred only in the latter. In conclusion, it is most striking that, although GPX1 protects against apoptosis induced by superoxide-generator DQ, the enzyme actually promotes apoptosis induced by PN in murine hepatocytes. Indeed, GSH is a physiological substrate for GPX1 in coping with ROS in these cells.
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PMID:Opposite roles of selenium-dependent glutathione peroxidase-1 in superoxide generator diquat- and peroxynitrite-induced apoptosis and signaling. 1156 67

Oxidants are known to induce cell apoptosis. Because oxidants also elicit redox imbalance, it is difficult to distinguish the direct effects of cellular redox from that of oxidants. This study tests the hypothesis that induction of redox imbalance independent of reactive oxygen species (ROS), can induce cell apoptosis in a mitotic competent, undifferentiated cell line, PC-12. Cells grown in standard DMEM containing 25 mM glucose were treated with diamide, a thiol oxidant, at a concentration that did not generate ROS. Diamide caused a rapid increase in oxidized glutathione (GSSG) and a loss of mitochondrial cytochrome c in 15-30 min, caspase-3 activation in 2 h, and apoptosis in 24 h. N-Acetyl cysteine attenuated GSSG elevation and diamide-induced apoptosis. Incubation of cells in 5 mM glucose or inhibition of the pentose phosphate pathway maintained GSSG elevation and accelerated cell apoptosis. Collectively, these results show that loss of redox balance is an upstream event that kinetically preceded mitochondrial apoptotic signaling. A sustained redox change was not critical or necessary for apoptotic progression, but its prolongation exacerbated apoptotic death. The potentiation of apoptosis by sustained redox imbalance was correlated with decreases in NADPH supply for GSSG reduction.
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PMID:Apoptosis in mitotic competent undifferentiated cells is induced by cellular redox imbalance independent of reactive oxygen species production. 1203 59


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