Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.22.56 (caspase-3)
 35,750 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The mechanisms by which immature thymocyte apoptosis is induced during negative selection are poorly defined. Reports demonstrated that cross-linking of T-cell receptor leads to stromal cell activation, expression of inducible nitric oxide synthase (iNOS) and, subsequently, to thymocyte apoptosis. Therefore we examined, whether NO directly or indirectly, through peroxynitrite formation, causes thymocyte apoptosis. Immuno-histochemical detection of nitrotyrosine revealed in vivo peroxynitrite formation in the thymi of naive mice. Nitrotyrosine, the footprint of peroxynitrite, was predominantly found in the corticomedullary junction and the medulla of naive mice. In the thymi of mice deficient in the inducible isoform of nitric oxide synthase, considerably less nitrotyrosine was found. Exposure of thymocytes in vitro to low concentrations (10 microM) of peroxynitrite led to apoptosis, whereas higher concentrations (50 microM) resulted in intense cell death with the characteristics of necrosis. We also investigated the effect of poly (ADP-ribose) synthetase (PARS) inhibition on thymocyte apoptosis. Using the PARS inhibitor 3-aminobenzamide (3-AB), or thymocytes from PARS-deficient animals, we established that PARS determines the fate of thymocyte death. Suppression of cellular ATP levels, and the cellular necrosis in response to peroxynitrite were prevented by PARS inhibition. Therefore, in the absence of PARS, cells are diverted towards the pathway of apoptotic cell death. Similar results were obtained with H2O2 treatment, while apoptosis induced by non-oxidative stimuli such as dexamethasone or anti-FAS antibody was unaffected by PARS inhibition. In conclusion, we propose that peroxynitrite-induced apoptosis may play a role in the process of thymocyte negative selection. Furthermore, we propose that the physiological role of PARS cleavage by apopain during apoptosis may serve as an energy-conserving step, enabling the cell to complete the process of apoptosis.
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PMID:Peroxynitrite-induced thymocyte apoptosis: the role of caspases and poly (ADP-ribose) synthetase (PARS) activation. 976 16

A novel anticancer drug, cytotrienin A, isolated from Streptomyces sp., induces apoptosis (or programmed cell death) in human promyelocytic leukemia HL-60 cells within 4 h. To elucidate the mechanism of this process, we performed an in-gel kinase assay using myelin basic protein (MBP) as a substrate and found the activation of kinase with an apparent molecular mass of 36 kDa (p36 MBP kinase). The dose of cytotrienin A required to activate p36 MBP kinase was consistent with that required to induce apoptotic DNA fragmentation in HL-60 cells. This p36 MBP kinase was activated with kinetics distinct from the activation of JNK (c-Jun N-terminal kinase)/stress-activated protein kinase and p38 MAPK (mitogen-activated protein kinase). Importantly, the p36 MBP kinase was immunologically different from MAPK superfamily molecules such as ERK1, JNK isoforms, and p38 MAPK. In addition, the p36 MBP kinase activation and apoptotic DNA fragmentation were inhibited by antioxidants such as N-acetylcysteine and reduced-form glutathione. The p36 MBP kinase activation was also observed during hydrogen peroxide (H2O2) and okadaic acid-induced apoptosis. Although a specific inhibitor of caspase-3-like proteases (Ac-DEVD-CHO) or a specific inhibitor of caspase-1-like proteases (Ac-YVAD-CHO) did not block the cytotrienin A-, H2O2-, or okadaic acid-induced apoptosis, a broad specificity inhibitor of caspases (Z-Asp-CH2-DCB) strongly inhibited the apoptosis of HL-60 cells. Surprisingly, Z-Asp-CH2-DCB inhibited the activation of p36 MBP kinase induced by cytotrienin A or H2O2, but did not inhibit the activation of JNK/stress-activated protein kinase and p38 MAPK. Taken together, these results indicate that p36 MBP kinase activation is downstream of the activation of Z-Asp-CH2-DCB-sensitive caspases, and reactive oxygen species could be included in the apoptotic events. Moreover, according to the Western blotting using the antibodies against MST1/Krs2 or MST2/Krs1, it is suggested that the p36 MBP kinase is an active proteolytic product of MST1/Krs2 and MST2/Krs1, which are originally cloned by virtue of its homology to the budding yeast Ste20 kinase. Thus, the p36 MBP kinase might be a common component of the diverse signaling pathways leading to apoptosis, and controlling this p36 MBP kinase pathway might be a novel strategy for cancer chemotherapy.
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PMID:Caspase-mediated activation of a 36-kDa myelin basic protein kinase during anticancer drug-induced apoptosis. 980 95

Pulsed field gel electrophoresis showed that the initiation time of DNA breakage induced by the DNA alkylating agent duocarmycin A, which is not a redox-cycling agent, was almost the same in the human leukemia cell line HL-60 and its H2O2-resistant clone HP100. Catalase activity of HP100 cells was much higher than that of HL-60 cells. Duocarmycin A-mediated DNA ladder formation in HP100 cells was delayed compared with that in HL-60 cells, suggesting the involvement of H2O2 in duocarmycin A-induced apoptosis. Flow cytometry demonstrated that peroxide formation preceded loss of mitochondrial membrane potential (delta psi m) in cells treated with duocarmycin A. Then, caspase-3 was activated, followed by DNA ladder formation. These findings suggest that DNA damage by duocarmycin A induces H2O2 generation, which causes delta psi m loss and subsequently caspase-3 activation, resulting in apoptosis.
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PMID:Generation of hydrogen peroxide precedes loss of mitochondrial membrane potential during DNA alkylation-induced apoptosis. 992 6

Beta-lapachone (beta-Lap) has been found to inhibit DNA topoisomerases (Topos) by a mechanism distinct from that of other commonly known Topo inhibitors. Here, we demonstrated a pronounced elevation of H2O2 and O2- in human leukemia HL-60 cells treated with beta-Lap. Treatment with other Topo poisons, such as camptothecin (CPT), Vbeta-16, and GL331, did not have the same effect. On the other hand, antioxidant vitamin C (Vit C) treatment effectively antagonized beta-Lap-induced apoptosis. This suggested that a reactive oxygen species (ROS)-related pathway was involved in beta-Lap-induced apoptosis program. We also found that c-Jun NH2-terminal kinase (JNK) but not p38 mitogen-activated protein kinase or extracellular signal-regulated kinase 1/2 was persistently activated in apoptosis induced by beta-Lap. Overexpression of a dominant-negative mutant mitogen-activated protein kinase kinase kinase 1 (MEKK1-DN) or treatment with JNK-specific antisense oligonucleotide or Vit C all prevented beta-Lap-induced JNK activation and the subsequent apoptosis. Only the expression of MEKK1-DN, not Vit C treatment, blocked the JNK activity induced by CPT, Vbeta-16, or GL331. These results confirm again that ROS acts as a mediator for JNK activation during beta-Lap-induced apoptosis. Furthermore, we found that beta-Lap can stimulate CPP32/Yama activity, which was, however, markedly inhibited by the MEKK1-DN expression or Vit C treatment. Again, CPT-induced CPP32/Yama activation can be abolished by MEKK1-DN but not by Vit C treatment. Taken together, these results indicate that beta-Lap but not other Topo inhibitors triggers apoptosis signaling, i.e., JNK and subsequent CPP32/Yama activation are mediated by the generation of ROS.
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PMID:Activation of c-Jun NH2-terminal kinase and subsequent CPP32/Yama during topoisomerase inhibitor beta-lapachone-induced apoptosis through an oxidation-dependent pathway. 992 52

LPS, a component of the cell wall in Gram-negative bacteria, induces inflammation and septic shock syndrome by stimulating various inflammatory cytokines including TNF. How LPS affects the TNF-mediated cellular responses, however, is not understood. In this study, the effect of LPS on TNF-mediated apoptosis in human histiocytic lymphoma U-937 cells was investigated. We found that treatment of cells with LPS completely abolished TNF-mediated cytotoxicity and activation of caspase-3. LPS-chelating antibiotic, polymyxin B, suppressed the antiapoptotic activity, indicating the specificity of the effect. Within minutes, LPS through CD14 induced the activation of NF-kappaB, degradation of IkappaBalpha (inhibitory subunit of NF-kappaB) and IkappaBbeta, and nuclear translocation of p65. An antioxidant, pyrrolidine dithiocarbamate, which blocked LPS-induced NF-kappaB activation, also abolished the antiapoptotic effects of LPS at the same time. Besides TNF, the apoptosis induced by taxol and okadaic acid was also sensitive to LPS-induced NF-kappaB activation, whereas that induced by H2O2, doxorubicin, daunomycin, vincristine, and vinblastine was NF-kappaB insensitive. Tumor cells that constitutively expressed NF-kappaB also showed resistance to the apoptotic effects of TNF, taxol, and okadaic acid, but sensitivity to all other agents, indicating the critical role of NF-kappaB in blocking apoptosis induced by certain agents. Overall, these results indicate that LPS induces resistance to the apoptotic effects of TNF and other agents, and that NF-kappaB activation, whether induced or constitutive, inhibits this apoptosis.
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PMID:Lipopolysaccharide inhibits TNF-induced apoptosis: role of nuclear factor-kappaB activation and reactive oxygen intermediates. 997 8

Recent studies have suggested that hydrogen peroxide (H2O2), a reactive compound formed endogenously in the breakdown of superoxide, may mediate the induction of apoptosis in various cell types in response to external stimuli. However, the role of H2O2 in the apoptotic pathway has not been clearly established. The purpose of this study was to determine if H2O2 treatment could induce apoptosis through the activation of caspases. Doses of H2O2 ranging from 10 microM to 100 microM, when added to HL-60 cells, resulted in the cleavage of poly(ADP-ribose) polymerase (PARP) from its native 113 Kd form to a processed 89 Kd fragment, indicative of cells undergoing apoptosis. PARP was predominantly in the fragmented form when doses of 20 microM and greater were used. A time course study of changes in PARP processing in H2O2-treated cells revealed that 10 and 50 microM H2O2 required 6 and 3 h, respectively, to specifically degrade PARP, suggesting that the H2O2-induced PARP cleavage is both time and concentration dependent. Since PARP is cleaved by CPP32 (caspase-3), we next determined if H2O2 was capable of effecting changes in CPP32 activity. The caspase activity was assayed using a colorimetric substrate, DEVD-pNa. Results of these experiments showed that H2O2 increased caspase activity at 3 h, corresponding to the time of appearance of fragmented PARP. Also, CPP32 activity and PARP processing were both significantly suppressed by caspase-3 inhibitors. Taken together, these results suggest that H2O2 mediates specific cleavage of PARP and possibly apoptosis by activating caspase 3.
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PMID:Activation of caspase 3 in HL-60 cells exposed to hydrogen peroxide. 1004 34

Beta-lapachone, the product of a tree from South America, is known to exhibit various pharmacologic properties, the mechanisms of which are poorly understood. In the present report, we examined the effect of beta-lapachone on the tumor necrosis factor (TNF)-induced activation of the nuclear transcription factors NF-kappaB and activator protein-1 (AP-1) in human myeloid U937 cells. TNF-induced NF-kappaB activation, p65 translocation, IkappaBalpha degradation, and NF-kappaB-dependent reporter gene expression were inhibited in cells pretreated with beta-lapachone. Direct treatment of the p50-p65 heterodimer of NF-kappaB with beta-lapachone had no effect on its ability to bind to the DNA. Besides myeloid cells, beta-lapachone was also inhibitory in T-cells and epithelial cells. Beta-lapachone also suppressed the activation of NF-kappaB by lipopolysaccharide, okadaic acid, and ceramide but had no significant effect on activation by H2O2 or phorbol myristate acetate, indicating that its action is selective. Beta-lapachone also abolished TNF-induced activation of AP-1, c-Jun N-terminal kinase, and mitogen-activated protein kinase kinase (MAPKK or MEK). TNF-induced cytotoxicity and activation of caspase-3 were also abolished by beta-lapachone. Because reducing agents (dithiothreitol and N-acetylcysteine) reversed the effect of beta-lapachone, it suggests the role of a critical sulfhydryl group. Overall, our results identify NF-kappaB, AP-1, and apoptosis as novel targets for beta-lapachone, and this may explain some of its pharmacologic effects.
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PMID:Suppression of tumor necrosis factor-activated nuclear transcription factor-kappaB, activator protein-1, c-Jun N-terminal kinase, and apoptosis by beta-lapachone. 1007 82

Protein phosphorylation in a human glioblastoma cell line, T98G, was examined after exposure to oxidative stress in vitro. Hydrogen peroxide (1 mM) markedly induced tyrosine phosphorylation of focal adhesion kinase (FAK) and serine phosphorylation of Akt at 1 h after stimulation. Concommitantly, the association of FAK with phosphatidylinositide 3'-OH-kinase (PI 3-kinase) was also observed by the hydrogen peroxide stimulation. When T98G cells were incubated with wortmannin, a PI 3-kinase inhibitor, both PI 3-kinase activity and phosphorylation of Akt were inhibited, whereas apoptosis by oxidative stress was accelerated. Concomitant with apoptosis, elevated level of CPP32 protease activity (caspase-3) was observed, with decreases in Bcl-2 protein and increases in Bax protein. These results suggested that in the signal transduction pathway from FAK to PI 3-kinase, Akt promotes survival. Thus, it became apparent that FAK is the upstream signal protein of the PI 3-kinase-Akt survival pathway in hydrogen peroxide-induced apoptosis in T98G cells.
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PMID:FAK is the upstream signal protein of the phosphatidylinositol 3-kinase-Akt survival pathway in hydrogen peroxide-induced apoptosis of a human glioblastoma cell line. 1018 51

Apoptosis has been associated with oxidative stress in biological systems. Caspases have been considered to play a pivotal role in the execution phase of apoptosis. However, which caspases function as executioners in reactive oxygen species (ROS)-induced apoptosis is not known. The present study was performed to identify the major caspases acting in ROS-induced apoptosis. Treatment of HL-60 cells with 50 microM hydrogen peroxide (H2O2) for 4 h induced the morphological changes such as condensed and/or fragmented nuclei, increase in caspase-3 subfamily protease activities, reduction of the procaspase-3 and a DNA fragmentation. To determine the role of caspases in H2O2-induced apoptosis, caspase inhibitors, acetyl-Tyr-Val-Ala-Asp-chloromethyl ketone (Ac-YVAD-cmk), acetyl-Asp-Glu-Val-Asp-aldehyde (Ac-DEVD-CHO) and acetyl-Val-Glu-Ile-Asp-aldehyde (Ac-VEID-CHO), selective for caspase-1 subfamily, caspase-3 subfamily and caspase-6, respectively, were loaded into the cells using an osmotic lysis of pinosomes method. Of these caspase inhibitors, only Ac-DEVD-CHO completely blocked morphological changes, caspase-3 subfamily protease activation and DNA ladder formation in H2O2-treated HL-60 cells. This inhibitory effect was dose-dependent. These results suggest that caspase-3, but not caspase-1 is required for commitment to ROS-triggered apoptosis.
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PMID:Hydrogen peroxide-induced apoptosis in HL-60 cells requires caspase-3 activation. 1019 75

Oxidative stress is considered to be an important pathophysiological condition to promote cell death in a broad variety of disorders, such as cardiovascular and neurodegenerative diseases. Scavestrogens, structurally derived from estradiol, are potent radical scavengers and inhibitors of iron-induced cell damage in vitro. In this study the potential cytoprotective effects of the so-called scavestrogen estra-1,3,5(10),8-tetraene-3,17alpha-diol, J 811, was tested using rat cerebellar granule cells (CGCs) exposed to 25 or 50 microM hydrogen peroxide (H2O2). H2O2-induced apoptotic cell death was detected by the appearance of high molecular weight DNA fragments and nuclear condensation. The addition of J 811 before or shortly after the exposure to H2O2 prevented CGC apoptosis in a dose-dependent manner. The estrogen receptor antagonist ICI 182.780 failed to prevent the protective effect of J 811, suggesting that the latter is not dependent on estrogen receptor activation. The lack of protection against apoptosis caused by colchicine suggests that J 811 is neither interfering with the activation of caspase-3, nor acting downstream of caspase-3. Therefore, the protective effect observed against H2O2 seems to be upstream caspases activation, pointing to a scavenging action of J 811. Thus the scavestrogen J 811 is a powerful antioxidant able to interfere with radical-mediated cell death and is potentially useful in diseases where reactive oxygen species are involved.
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PMID:Radical scavenging compound J 811 inhibits hydrogen peroxide-induced death of cerebellar granule cells. 1034 Jul 49


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