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)

Oxygen deprivation for prolonged periods leads to cardiac cell death and ventricular dysfunction. The ability to prevent myocardial cell death would be of significant therapeutic value in maintaining cardiac function after injury. While caspases have been suggested to play a critical role in apoptosis, their involvement during hypoxic injury has not been formally determined. In this report, we show that adult ventricular myocytes subjected to hypoxia for 1 h undergo a three-fold increase (P<0.05) in the incidence of apoptosis as determined by TUNEL analysis and Hoechst 33258 nuclear staining. Western blot analysis of hypoxic myocytes revealed a 10-fold increase in the proteolytic processing of caspase 3 to p17 with a concomitant cleavage of the caspase 3 substrate PARP from 116 kd to p85 kd compared to normoxic controls. Defects in mitochondrial membrane integrity were also observed as evidenced by the translocation of cytochrome c from the mitochondrial to cytosolic compartment of hypoxic cells. Pretreatment of ventricular myocytes with the peptide-caspase inhibitor known to block caspases related to caspase 1 (Ac-YVAD-CHO) attenuated cytochrome c release, processing of caspase 3, and apoptosis. While the caspase inhibitor (Ac-DEVD-CHO) which blocks caspases related to caspase 3, suppressed the cleavage of PARP and apoptosis, it had no effect on cytochrome c release by mitochondria. The data provide direct evidence for the proteolytic activation of caspases during hypoxia-mediated apoptosis of adult ventricular myocytes. Furthermore, the data suggest a hierarchical scheme for caspase activation with mitochondrial cytochrome c release occurring proximally to DEVD-CHO-inhibitable caspases.
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PMID:Caspase activation and mitochondrial cytochrome C release during hypoxia-mediated apoptosis of adult ventricular myocytes. 1065 90

Nordihydroguaiaretic acid (NDGA) induces apoptosis in a variety of cell lines. The mechanism(s) of this effect is not known, although the focus has been on the ability of NDGA to inhibit lipoxygenase (LOX) activities. In the present study, NDGA-induced apoptosis was studied in a murine hematopoietic cell line, FL5.12. Although this cell line lacks detectable LOX protein or activities, NDGA (10 microM) was able to induce apoptosis. There was a massive loss of mitochondrial membrane potential by 4 h after the addition of NDGA, suggesting that this organelle might be targeted by NDGA. A pro-oxidant NDGA effect has been suggested as playing a role in apoptosis. This was supported by the findings that glutathione disulfide levels were increased by 4 h following treatment with 10 microM NDGA, that pretreatment with N-acetylcysteine completely blocked the NDGA-induced loss of membrane potential and apoptosis, and that lipid peroxidation was enhanced in cells treated with NDGA. However, no evidence of increased levels of reactive oxygen could be seen in NDGA-treated cells loaded with dichlorofluorescin diacetate or dihydrorhodamine and analyzed by flow cytometry. Bcl-X(L) protein levels were unaffected by NDGA treatment. Caspase-3 was rapidly activated with a peak at 8 h after FL5.12 cells were treated with NDGA. Ac-DEVD-CHO (25 microM) and boc-asp-FMK (20 microM) both inhibited caspase-3 enzyme activity by 97% 8 h after NDGA treatment. Boc-asp-FMK, a more general caspase inhibitor, delayed NDGA-induced apoptosis while Ac-DEVD-CHO, a more specific inhibitor of caspase-3, had no effect. These results suggest that NDGA-induced apoptosis happens through reactions that depolarize mitochondria, oxidize glutathione and lipids, but do not generate significant amounts of free reactive oxygen species.
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PMID:Glutathione oxidation and mitochondrial depolarization as mechanisms of nordihydroguaiaretic acid-induced apoptosis in lipoxygenase-deficient FL5.12 cells. 1065 24

Nitric oxide (NO) challenge to human neuroblastoma cells (SH-SY5Y) ultimately results in apoptosis. Tumor suppressor protein p53 and cell cycle inhibitor p21 accumulate as an early sign of S-nitrosoglutathione-mediated toxicity. Cytochrome c release from mitochondria and caspase 3 activation also occurred. Cells transfected with either wild type (WT) or mutant (G93A) Cu, Zn-superoxide dismutase (Cu,Zn-SOD) produced comparable amounts of nitrite/nitrate but showed different degree of apoptosis. G93A cells were the most affected and WT cells the most protected; however, Cu, Zn-SOD content of these two cell lines was 2-fold the SH-SY5Y cells under both resting and treated conditions. We linked decreased susceptibility of the WT cells to higher and more stable Bcl-2 and decreased reactive oxygen species. Conversely, we linked G93A susceptibility to increased reactive oxygen species production since simultaneous administration of S-nitrosoglutathione and copper chelators protects from apoptosis. Furthermore, G93A cells showed a significant decrease of Bcl-2 expression and, as target of NO-derived radicals, showed lower cytochrome c oxidase activity. These results demonstrate that resistance to NO-mediated apoptosis is strictly related to the level and integrity of Cu,Zn-SOD and that the balance between reactive nitrogen and reactive oxygen species regulates neuroblastoma apoptosis.
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PMID:Cu,Zn-superoxide dismutase-dependent apoptosis induced by nitric oxide in neuronal cells. 1067 49

The aim of this study was to investigate the mechanism of flavonoid-induced apoptosis in HL-60 leukaemic cells. Thus, the effect of structurally related flavonoids on cell viability, DNA fragmentation and caspase activity was assessed. Loss of membrane potential and reactive oxygen species generation were also monitored by flow cytometry. The structurally related flavonoids, such as apigenin, quercetin, myricetin, and kaempferol were able to induce apoptosis in human leukaemia HL-60 cells. Treatment with flavonoids (60 microM) caused a rapid induction of caspase-3 activity and stimulated proteolytic cleavage of poly-(ADP-ribose) polymerase (PARP). Furthermore, these flavonoids induced loss of mitochondrial transmembrane potential, elevation of reactive oxygen species (ROS) production, release of mitochondrial cytochrome c into the cytosol, and subsequent induction of procaspase-9 processing. The potency of these flavonoids on these features of apoptosis were in the order of: apigenin > quercetin > myricetin > kaempferol in HL-60 cells treated with 60 microM flavonoids. These results suggest that flavonoid-induced apoptosis is stimulated by the release of cytochrome c to the cytosol, by procaspase-9 processing, and through a caspase-3-dependent mechanism. The induction of apoptosis by flavonoids may be attributed to their cancer chemopreventive activity. Furthermore, the potency of flavonoids for inducing apoptosis may be dependent on the numbers of hydroxyl groups in the 2-phenyl group and on the absence of the 3-hydroxyl group. This provides new information on the structure-activity relationship of flavonoids.
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PMID:Induction of apoptosis by apigenin and related flavonoids through cytochrome c release and activation of caspase-9 and caspase-3 in leukaemia HL-60 cells. 1067 81

Glial cell line-derived neurotrophic factor (GDNF) provides neuroprotection, but its neuroprotective mechanism has not been resolved. We investigated the neuroprotective mechanism of GDNF using primary culture of the rat mesencephalon. Bleomycin sulfate (BLM) and L-buthionine-[S,R]-sulfoximine (BSO) caused apoptosis in both dopaminergic and nondopaminergic neurons, as revealed by the presence of chromatin condensation, and positive staining by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL). GDNF preincubation blocked the neurotoxicity and reduced the number of the TUNEL-positive cells caused by BLM and BSO exposure. In contrast, GDNF did not provide neuroprotection against glutamate toxicity, which was not accompanied by these apoptotic features. The neuroprotection was mediated by phosphatidylinositol 3-kinase, an effector downstream from c-Ret, because it was blocked by LY294002. GDNF pretreatment caused up-regulation of Bcl-2 and Bcl-x. Furthermore, GDNF suppressed oxygen radical accumulation caused by BLM. Apoptosis induced by BLM and BSO was blocked by a caspase-3 inhibitor. Caspase-3 activity was elevated by BLM and suppressed by GDNF pretreatment. These findings indicate that GDNF has no effect on necrosis but exerts protection against apoptosis by activation of phosphatidylinositol 3-kinase and the subsequent up-regulation of Bcl-2 and Bcl-x, which suppresses accumulation of oxygen radicals followed by caspase-3 activation.
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PMID:Neuroprotective mechanism of glial cell line-derived neurotrophic factor in mesencephalic neurons. 1069 50

Interleukin 8 (IL-8) and growth-related oncogene alpha (Gro-alpha) delay neutrophil apoptosis, which is thought to be important for the resolution of inflammation. We hypothesized that (IL-8) and Gro-alpha interfere with extracellular death receptor signaling or intracellular caspase activation to suppress neutrophil apoptosis. In addition, we sought to determine if prolonged neutrophil half-life was associated with preservation of function. Polymorphonuclear leukocytes (PMN) were cultured with IL-8 or Gro-alpha (0-100 ng/mL) in normoxia or hypoxia, and the extent of apoptosis was assessed by histology and TdT-mediated dUTP nick end labeling (TUNEL). Subsequently, to determine the role of apoptotic-associated receptors, PMN were cultured with IL-8 and neutralizing monoclonal antibody to Fas (CD95), TNFR55, and TNFR75. To establish the effect of IL-8 or Gro-alpha on pro-apoptotic caspase activity, the cleavage of specific colorimetric substrates was assessed. Functional changes in PMN included the capacity to produce superoxide anion and phagocytosis of Escherichia coli. At the 100 ng/mL dose, the addition of IL-8 and Gro-alpha maximally suppressed PMN apoptosis from 54% (untreated) to 5% and 6%, respectively. The addition of neutralizing antibodies to Fas, TNFR55, or R75 caused no change in IL-8 suppression of apoptosis. Caspase 3 activity was markedly suppressed at 24 h by the inclusion of either IL-8 and Gro-alpha. IL-8 and Gro-alpha-stimulated PMN released more superoxide anion and had an increased phagocytic index vs. control PMN. IL-8 and Gro-alpha suppress neutrophil apoptosis to a similar level that is not influenced by oxygen tension at high doses. The effect of IL-8 and Gro-alpha does not depend on activation of the Fas, TNFR55, or R75 receptor pathways but involves suppression of caspase 3 activity. IL-8 or Gro-alpha extends the functional half-life of neutrophils and may explain their role in disease states such as acute respiratory distress syndrome.
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PMID:CXC chemokine suppression of polymorphonuclear leukocytes apoptosis and preservation of function is oxidative stress independent. 1071 83

Some, but not all, of a series of novel pyrrolo-1,5-benzoxazepines (PBOXs) induce apoptosis as shown by cell shrinkage, chromatin condensation, and DNA fragmentation in three human cell lines, HL-60 promyelocytic, Jurkat T lymphoma, and Hut-78 s.c. lymphoma cells. This chemical selectivity, together with the lack of apoptotic activity against rat Leydig cells, argues against a general cell poisoning effect. PBOX-6, a potent member of the series, caused activation of a member of the caspase-3 family of proteases. In addition, the caspase-3-like inhibitor z-DEVD-fmk, but not the caspase-1-like inhibitor z-YVAD-fmk prevented PBOX-6-induced apoptosis, suggesting that caspase 3-like proteases are involved in the mechanism by which PBOX compounds induce apoptosis. The release of cytochrome c into the cytosol in HL-60 cells in response to PBOX-6 suggests that this cellular response may be important in the mechanism by which PBOX-6 induces apoptosis. However, reactive oxygen intermediates do not play a key role in PBOX-6-induced apoptosis because neither the free radical scavenger TEMPO nor the antioxidant N-acetylcysteine had any effect on PBOX-6-induced apoptosis. The apoptotic induction seems independent of the mitochondrial peripheral-type benzodiazepine receptor (PBR) that binds these pyrrolobenzoxazepines with high affinity, due to the lack of correlation between their affinities for the receptor and their apoptotic potencies, their high apoptotic activity in PBR-deficient cells such as Jurkats, and their lack of apoptotic induction in PBR-rich rat Leydig cells. These PBOXs also can overcome nuclear factor-kappaB-mediated resistance to apoptosis. This suggests an important potential use of these compounds in drug-resistant cancers.
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PMID:Pyrrolo-1,5-benzoxazepines induce apoptosis in HL-60, Jurkat, and Hut-78 cells: a new class of apoptotic agents. 1073 52

The induction of cell death in leukemic HL-60 cells by the ether lipid 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH(3); edelfosine) followed the typical apoptotic changes in ultrastructural morphology, including blebbing, chromatin condensation, nuclear membrane breakdown and extensive vacuolation. Using a cytofluorimetric approach, we found that ET-18-OCH(3) induced disruption of the mitochondrial transmembrane potential (DeltaPsi(m)) followed by production of reactive oxygen species (ROS) and DNA fragmentation in leukemic cells. ET-18-OCH(3) also induced caspase-3 activation in human leukemic cells, as assessed by cleavage of caspase-3 into the p17 active form and cleavage of the caspase-3 substrate poly(ADP-ribose) polymerase (PARP). ET-18-OCH(3) analogues unable to induce apoptosis failed to disrupt DeltaPsi(m) and to activate caspase-3. ET-18-OCH(3)-resistant Jurkat cells generated from sensitive Jurkat cells showed no caspase-3 activation and did not undergo DeltaPsi(m) disruption upon ET-18-OCH(3) incubation. Cyclosporin A partially inhibited DeltaPsi(m) dissipation, caspase activation and apoptosis in ET-18-OCH(3)-treated leukemic cells. Overexpression of bcl-2 by gene transfer prevented DeltaPsi(m) collapse, ROS generation, caspase activation and apoptosis in ET-18-OCH(3)-treated leukemic T cells. Pretreatment with the caspase inhibitor Z-Asp-2, 6-dichlorobenzoyloxymethylketone prevented ET-18-OCH(3)-induced PARP proteolysis and DNA fragmentation, but not DeltaPsi(m) dissipation. ET-18-OCH(3) did not affect the expression of caspases and bcl-2-related genes. ET-18-OCH(3)-induced apoptosis did not require protein synthesis. Our data indicate that DeltaPsi(m) dissipation and caspase-3 activation are critical events of the apoptotic cascade triggered by the antitumor ether lipid ET-18-OCH(3), and that the sequence of events in the apoptotic action of ET-18-OCH(3) on human leukemic cells is: DeltaPsi(m) disruption, caspase-3 activation and internucleosomal DNA degradation.
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PMID:Involvement of mitochondria and caspase-3 in ET-18-OCH(3)-induced apoptosis of human leukemic cells. 1073 48

Apoptosis, or programmed cell death, plays a central role in the development and homeostasis of an organism. The breakdown of cellular proteins in apoptosis is mediated by caspases, which comprise a highly conserved family of cysteine proteases with specificity for aspartic acid residues at the P1 positions of their substrates. Multiple lines of evidence show that caspase-9 is critical for an apoptosis pathway mediated via the mitochondria. In this study, the three-dimensional structure of the catalytic domain of caspase-9 and its interaction with the inhibitor acetyl-Asp-Val-Ala-Asp fluoromethyl ketone (Ac-DVAD-fmk) have been predicted by a segment matching modeling procedure. As expected, the predicted caspase-9 structure shows both a high similarity in the overall folding topology and remarkable differences in the surface loop regions as compared to other caspase family members such as caspase-1, -3 and -8, for which crystal structures have been determined. This kind of comparative analysis reflects the convergence-divergence duality among the caspases. Moreover, some subtle differences have been observed between caspase-9 and caspase-3 in the subsite contacts with the covalently linked inhibitor Ac-DVAD-fmk. Based on the X-ray structural analysis of caspase-8, a main chain carbonyl oxygen appears to be involved in a catalytic triad with the active site Cys and His residues. The corresponding carbonyl oxygen in caspase-9, together with other expected features of the catalytic apparatus, appears in our model. The predicted structure of caspase-9 can serve as a reference for subsite analysis relative to rational design of highly selective caspase inhibitors for therapeutic application.
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PMID:Prediction of the tertiary structure of a caspase-9/inhibitor complex. 1074 77

Renal failure associated with aspergillosis is caused by pathogenic fungi. Gliotoxin is a toxic epipolythiodioxopiperazine metabolite produced by the pathogens. The present study investigated the cytotoxicity and underlying mechanisms induced by gliotoxin in LLC-PK1 cells, a porcine renal proximal tubular cell line. Gliotoxin at 100 ng/ml did not show a cytotoxic effect, but unmasked a dose-dependent cell death induced by TNF-alpha. TNF-alpha-induced cell death in the presence of gliotoxin was associated with hypodiploid nuclei and activation of caspase-3-like proteases. Blockade of caspases by boc-aspartyl (OMe)-fluoromethylketone and z-DEVD.fmk inhibited TNF-alpha-induced cell death. As the concentrations of gliotoxin were increased, gliotoxin killed the cells directly in a dose-dependent manner. Further analyses of DNA fragmentation, hypodiploid nuclei, mitochondrial membrane potential, and plasma membrane integrity revealed that cell death proceeded via apoptosis. Gliotoxin-induced apoptosis was associated with dose-dependent and time-dependent activation of caspase-3-like proteases. Boc-aspartyl (OMe)-fluoromethylketone attenuated the killing effect. Gliotoxin also increased the intracellular levels of reactive oxygen species as measured by flow cytometry. N-acetylcysteine, a well-known antioxidant, completely abolished the gliotoxin-induced caspase-3-like activity, cytotoxicity, and reactive oxygen species. In conclusion, (1) gliotoxin at 100 ng/ml unmasks the ability of TNF-alpha-induced apoptosis, and the effect of TNF-alpha is mediated by caspase-3-like proteases; and (2) at higher concentrations gliotoxin itself induces cell death, which is via apoptosis and dependent on caspase-3-like activity and reactive oxygen species.
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PMID:Gliotoxin-induced cytotoxicity proceeds via apoptosis and is mediated by caspases and reactive oxygen species in LLC-PK1 cells. 1074 46


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