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 inhibitory effect of nitric oxide (NO) on the enzymatic activity of CPP32-like proteases in the cell extract from vincristine-treated cells was examined in vitro. NO generated from (+/-)-(E)-methyl-2-[(E)-hydroxyimino]-5-nitro-6-methoxy-S-hexen eamide (NOR1) inhibited CPP32-like protease, which constitute a family of interleukin-1beta-converting enzyme (ICE)-like proteases in a dose-dependent manner. Moreover, recombinant CPP32beta activity was inhibited by NOR1 at same concentration. Inhibition of CPP32-like activity by NO was reversed in the presence of glutathione in the enzymatic reaction mixture. Thus, CPP32-like activity was regulated by NO under redox regulation. These findings suggest that NO may prevent apoptosis by inhibiting the ICE protease cascade under the influence of cellular redox status.
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PMID:Nitric oxide inhibits CPP32-like activity under redox regulation. 924 Apr 42

Nitric oxide (NO) generated from 1-hydroxy-2-oxo-3, 3-bis(2-aminoethyl)-1-triazene (NOC 18), an NO-releasing compound, induced monocytic differentiation of human promyelocytic leukemia HL-60 cells as assessed by expression of nonspecific esterases and morphologic maturation. Simultaneously, DNA fragmentation and morphological alterations typical of apoptosis were also induced. To investigate the mechanisms of apoptosis during differentiation of HL-60 cells induced by NO, the endogenous levels of Bcl-2 and Bax were assessed by immunoblotting. Treatment of cells with NOC 18 slightly reduced the level of Bcl-2 followed by Bax. These changes might be involved in the induction of apoptosis. The involvement of the activation of the interleukin-1 beta converting enzyme (ICE) family of proteases (caspases), such as ICE and CPP32, in the pathways was also investigated. CPP32, but not ICE, was strongly activated in response to NOC 18 stimulation, thereby implicating CPP32-like activity in the induction of apoptosis. Moreover, the possible involvement of tyrosine phosphorylation in apoptosis was investigated. Pretreatment of cells with herbimycin A, an inhibitor of tyrosine kinases, suppressed DNA fragmentation and CPP32-like activity, whereas pretreatment with vanadate, an inhibitor of tyrosine phosphatases, enhanced both parameters, suggesting that tyrosine phosphorylation might be involved in the pathways of apoptosis in HL-60 cells induced by NO.
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PMID:Molecular mechanisms of apoptosis in HL-60 cells induced by a nitric oxide-releasing compound. 935 Apr 36

Nitric oxide (NO) has emerged as an important endogenous inhibitor of apoptosis, and here we report that NO prevents hepatocyte apoptosis initiated by the removal of growth factors or exposure to TNFalpha or anti-Fas antibody. We postulated that the mechanism of the inhibition of apoptosis by NO would include an effect on caspase-3-like protease activity. Caspase-3-like activity increased coincident with apoptosis due to all three stimuli, and treatment with the caspase-3-like protease inhibitor N-acetyl-Asp-Glu-Val-Asp-aldehyde inhibited both proteolytic activity and apoptosis. Endogenous or exogenous sources of NO prevented the increase in caspase-3-like activity in hepatocytes. Exposure of purified recombinant caspase-3 to an NO or NO+ donor inhibited proteolytic activity. Dithiothreitol (DTT), but not glutathione, reversed the inhibition of recombinant caspase-3 by NO. When lysates from cells stimulated to express inducible NO synthase or cells exposed to NO donors were incubated in DTT, caspase-3-like activity increased to about 55% of cells not exposed to a source of NO. Similarly, administration of an NO donor to rats treated with TNFalpha and D-galactosamine also prevented the increase in caspase-3-like activity as measured in liver homogenates. The effect of the NO donor was reversed by about 50% if the homogenate was incubated with DTT. TNFalpha-induced apoptosis and caspase-3-like activity were also reduced in cultured hepatocytes exposed to 8-bromo-cGMP, and both effects were inhibited by the cGMP-dependent kinase inhibitor KT5823. The suppression in caspase-3-like activity in hepatocytes exposed to an NO donor was partially blocked by an inhibitor of soluble guanylyl cyclase, 1H-[1,2,4]oxadiazolo[4,3, -a]quinoxalin-1-one, (ODQ), while the incubation of these lysates in DTT almost completely restored caspase-3-like activity to the level of TNFalpha-treated controls. These data indicate that NO prevents apoptosis in hepatocytes by either directly or indirectly inhibiting caspase-3-like activation via a cGMP-dependent mechanism and by direct inhibition of caspase-3-like activity through protein S-nitrosylation.
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PMID:Nitric oxide inhibits apoptosis by preventing increases in caspase-3-like activity via two distinct mechanisms. 938 67

Poly(ADP-ribose)polymerase (PARP, EC 2.4.2.30), an abundant nuclear protein activated by DNA nicks, mediates cell death in vitro by nicotinamide adenine dinucleotide (NAD) depletion after exposure to nitric oxide. The authors examined whether genetic deletion of PARP (PARP null mice) or its pharmacologic inhibition by 3-aminobenzamide (3-AB) attenuates tissue injury after transient cerebral ischemia. Twenty-two hours after reperfusion following 2 hours of filamentous middle cerebral artery occlusion, ischemic injury was decreased in PARP-/- and PARP+/- mice compared with PARP+/+ litter mates, and also was attenuated in 129/SV wild-type mice after 3-AB treatment compared with controls. Infarct sparing was accompanied by functional recovery in PARP-/- and 3-AB-treated mice. Increased poly(ADP-ribose) immunostaining observed in ischemic cell nuclei 5 minutes after reperfusion was reduced by 3-AB treatment. Levels of NAD--the substrate of PARP--were reduced 2 hours after reperfusion and were 35% of contralateral levels at 24 hours. The decreases were attenuated in PARP-/- mice and in 3-AB-treated animals. Poly(ADP-ribose)polymerase cleavage by caspase-3 (CPP-32) has been proposed as an important step in apoptotic cell death. Markers of apoptosis, such as oligonucleosomal DNA damage, total DNA fragmentation, and the density of terminal deoxynucleotidyl transferase dUTP nick-end-labelled (TUNEL +) cells, however, did not differ in ischemic brain tissue of PARP-/- mice or in 3-AB-treated animals versus controls, although there were differences in the number of TUNEL-stained cells reflecting the decrease in infarct size. Thus, ischemic brain injury activates PARP and contributes to cell death most likely by NAD depletion and energy failure, although the authors have not excluded a role for PARP in apoptotic cell death at earlier or later stages in ischemic cell death. Inhibitors of PARP activation could provide a potential therapy in acute stroke.
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PMID:Ischemic brain injury is mediated by the activation of poly(ADP-ribose)polymerase. 939 Jun 45

Physiological levels of shear stress reduce endothelial cell turnover and exert a potent antiatherosclerotic effect. Here we demonstrate that oxidative stress-induced apoptosis of human endothelial cells was inhibited by shear stress exposure (15 dynes/cm2). Incubation with H2O2 (200 mumol/L) for 18 hours induced apoptosis of human umbilical venous endothelial cells as demonstrated by an enzyme-linked immunosorbent assay specific for histone-associated DNA fragments and visual analysis of fluorescence-stained nuclei. Shear stress-mediated inhibition of apoptosis was partially prevented by pharmacological inhibition of glutathione (GSH) biosynthesis with buthionine sulfoximine (BSO) or nitric oxide (NO) synthase with NG-monomethyl-L-arginine (LNMA), whereas inhibition of catalase by aminotriazol did not affect the inhibitory action of shear stress. Combined inhibition of NO synthase and GSH biosynthesis completely reversed the protective effect of shear stress, suggesting that both NO synthase and the GSH redox cycle system are involved in the apoptosis-suppressing effect of shear stress. Similar results were obtained when apoptosis was stimulated by tumor necrosis factor alpha (TNF alpha). To gain further insights into the interference of shear stress with apoptosis signal transduction, we measured caspase-3-like activity, a cysteine protease that has been shown to play a predominant role in the cell death effector pathway. Indeed, shear stress prevented the activation of caspase-3-like activity induced by H202 or TNF alpha. The inhibitory effect of shear stress was prevented by LNMA and BSO, suggesting that the reduction of oxidative flux by shear stress prevents the activation of caspase-like proteases and thereby inhibits apoptotic cell death in human endothelial cells.
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PMID:Shear stress inhibits H2O2-induced apoptosis of human endothelial cells by modulation of the glutathione redox cycle and nitric oxide synthase. 943 9

Nitric oxide has been shown to inhibit apoptosis of human umbilical venous endothelial cells (HUVEC). Therefore we investigated the effect of different NO donors, PAPA NONOate (NOC-15; NO.) and nitrosodium tetrafluoroborate (NOBF4, NO+), and the reaction product of NO and O2-, peroxynitrite (ONOO- ), on TNF-alpha- or serum depletion-induced apoptosis of HUVEC. TNF-alpha-induced DNA fragmentation, determined by ELISA, was inhibited by NOC-15, NOBF4, and ONOO- in a concentration-dependent manner (maximal effects with 10 microM NO. and ONOO- and 100 microM NO+). The inhibition of apoptosis correlated with a protective effect on cell viability. The caspases, a cysteine protease family, play an important role in apoptotic processes. To determine whether the different NO donors and ONOO- regulate this enzyme, caspase-3-like activity was measured in homogenates of TNF-alpha-treated HUVEC. The TNF-alpha-induced enzyme activity was abrogated by NO., NO+, and ONOO-. Furthermore, caspase-3 activity was determined in vitro by reconstitution of the separately cloned, bacterially expressed, and purified active p17 and p12 subunits. The reconstituted caspase-3 exhibited enzyme activity, which was suppressed by the different NO donors and ONOO- with an IC50 of 50 microM for NOC-15, 1 mM for NOBF4, and 50 microM for ONOO-. The inhibition of caspase-3 activity correlated with a S-nitrosylation of the reactive cysteine residue and was reversed by further addition of dithiothreitol. This study suggests that the cellular regulatory processes of NO to protect cells from apoptosis may be independent of the redox state and that low concentrations of NO and ONOO- inhibit the cellular suicide program in HUVEC via S-nitrosylation of members of the caspase family.
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PMID:Effects of redox-related congeners of NO on apoptosis and caspase-3 activity. 944

The reaction of superoxide and nitric oxide results in the formation of peroxynitrite, a long lived and highly reactive oxidant species. It has been suggested that the formation of peroxynitrite in vivo may contribute to cell death in some neurological conditions. We have examined the effect of peroxynitrite on cell death in the NSC34 spinal cord cell line. A brief (30 min) exposure to either peroxynitrite or hydrogen peroxide caused delayed cell death with an EC50 for both of approximately 1 mM. Cell death was prevented by the RNA synthesis inhibitor actinomycin D and included DNA damage as an early event. We sought to clarify the potential role of the DNA binding enzyme poly(ADP-ribose) polymerase (PARP) in cell death in these cells. Several PARP inhibitors [benzamide, 3-aminobenzamide, nicotinamide, and 6(5H)-phenanthridinone] prevented cell death, but the inactive analogue benzoic acid did not. However, there was no evidence of cleavage of PARP, which occurs in apoptosis via the activation of the caspase CPP32. Therefore, we suggest that PARP contributes to neuronal injury as an early event, probably by lethal NAD depletion, without any requirement for proteolytic cleavage.
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PMID:Peroxynitrite and hydrogen peroxide induced cell death in the NSC34 neuroblastoma x spinal cord cell line: role of poly (ADP-ribose) polymerase. 945 43

Previously we have shown that nitric oxide (NO) donors induced apoptosis in vascular smooth muscle cells (VSMCs). However, the mechanisms by which NO induced apoptosis in VSMCs are entirely unknown. In the present study, we intended to identify the mechanism by which NO donors induce apoptosis in VSMCs. First, we evaluated the expression of c-Myc, P53, and Bcl-2 proteins in VSMCs treated by NO donors. c-Myc and P53 protein expression increased after VSMCs were incubated with NO donors for 6 hr and reached a maximum level at 24 hr, while Bcl-2 protein decreased after 12 hr incubation. Next we investigated to see whether the CPP32 protease activation was involved in NO donors-induced apoptosis. In VSMCs treated by NO donors, the increase of CPP32 protease activity was observed and specific inhibition of CPP32 activity significantly prevented apoptosis induced by NO donors in a dose-dependent manner. These results suggest that NO donors induced apoptosis through proto-oncoprotein expression and CPP32-like protease activation.
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PMID:No induced apoptosis accompanying the change of oncoprotein expression and the activation of CPP32 protease. 948 2

The formation of nitric oxide (NO.) and superoxide (O2-) promotes rat mesangial cell death. Apoptotic death is characterized by DNA fragmentation, caspase-3 activation and concomitant poly(ADPribose) polymerase cleavage, as well as accumulation of the tumor suppressor protein p53. In close association with apoptotic parameters we noticed upregulation of heme oxygenase by the NO donor S-nitrosoglutathione (GSNO) and the redox cycler 2,3-dimethoxy-1,4-naphthoquinone (DMNQ) in a time- and concentration-dependent manner. In response to the NO. donor, heme oxygenase-1 expression was more easily obtained than initiation of apoptosis. Radical (NO./O2-) cogeneration abrogated DNA fragmentation, suppressed caspase activation and lowered p53 accumulation, thereby promoting cell survival of mesangial cells. In contrast, heme oxygenase-1 expression remained elevated under conditions of GSNO/DMNQ coadministration. Conclusively, heme oxygenase-1 is a stress marker for both nitrosative and oxidative stress. Accumulation of heme oxygenase-1 is found under conditions of both, apoptotic cell death and cell survival, thereby questioning a specific cytoprotective role of heme oxygenase-1 under conditions of NO. and/or O2- formation in rat mesangial cells.
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PMID:Nitrosative and oxidative stress induced heme oxygenase-1 accumulation in rat mesangial cells. 954 95

Apoptosis is an active process critical for the homeostasis of organisms. Enzymes of the caspase family are responsible for executing this process. We have previously shown that peroxynitrite (ONOO-), a biological product generated from the interaction of nitric oxide and superoxide, induces apoptosis of HL-60 cells. The aim of this study was to elucidate the mechanisms involved in the execution process of peroxynitrite-induced apoptosis. Proteolytic cleavage of poly(ADP-ribose) polymerase, an indication of caspase-3 family protease activation and an early biochemical event accompanying apoptosis, was observed in a time-dependent manner during peroxynitrite-induced apoptosis of HL-60 cells. Activation of caspase-3 during peroxynitrite-induced apoptosis was substantiated by monitoring proteolysis of the caspase-3 proenzyme and by measuring caspase-3 activity with a fluorogenic substrate. Furthermore, pretreatment of HL-60 cells with N-acetyl-Asp-Glu-Val-Asp-aldehyde, a specific inhibitor of caspase-3, but not N-acetyl-Tyr-Val-Ala-Asp-aldehyde, a specific inhibitor of caspase-1, decreased peroxynitrite-induced apoptosis. These results suggest that the activation of a caspase-3 family protease is essential for initiating the execution process of peroxynitrite-induced apoptosis of HL-60 cells.
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PMID:Peroxynitrite induces apoptosis of HL-60 cells by activation of a caspase-3 family protease. 957 79


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