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

Cyclooxygenase-2 (Cox-2), an enzyme responsible for catalyzing the committed step in prostanoid biosynthesis, is the product of an immediate early gene capable of being up-regulated by diverse stimuli. Significantly Cox-2 mRNA is absent from rat pheochromocytoma (PC12) cells, both basally and following stimulation with a range of agonists. Using PC12 cells engineered to stably express isopropyl-1-thio-beta-D-galactopyranoside-inducible Cox-2 (PCXII-4), we have investigated the putative effects of Cox-2 expression on differentiation, proliferation, and trophic withdrawal apoptosis. Cox-2 bioactivity had no effect on nerve growth factor-induced differentiation, epidermal growth factor-induced proliferation, or aromatic L-amino acid decarboxylase expression. However, trophic withdrawal apoptosis, induced by the removal of nerve growth factor following differentiation, was markedly reduced in the PCXII-4 when compared with control cells, as assessed by annexin V staining, DNA laddering, and Hoechst 33258 staining. The specificity of this effect was confirmed using two pharmacologically distinct nonsteroidal anti-inflammatory drugs, indomethacin and NS398. Investigations showed that the activity of the pro-apoptotic protease caspase-3 was reduced in PCXII cells. This study demonstrates that Cox-2-derived prostaglandins exert cytoprotective effects in trophic factor withdrawal apoptosis and provides evidence that this is, at least in part, due to suppression of caspase-3 activity.
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PMID:Cyclooxygenase-2 expression inhibits trophic withdrawal apoptosis in nerve growth factor-differentiated PC12 cells. 1076 43

Apoptosis is a process of active cell death and is characterized by activation of caspases, DNA fragmentation, and biochemical and morphological changes. To better understand apoptosis, we have characterized the dose- and time-dependent toxic effects of cadmium in Rat-1 fibroblasts. Staining of cells with phosphatidylserine (PS)-annexin V, Hoechst 33258 or Rhodamine 123 and Tunel assays showed that incubating cells with 10 microM cadmium induced a form of cell death exhibiting typical characteristics of apoptosis, including cell shrinkage, externalization of PS, loss of mitochondria membrane potential, nuclear condensation and DNA fragmentation. Expression of Bcl-2 or CrmA each suppressed cadmium-induced cell death although Bcl-2 was somewhat more effective than CrmA. In vitro assay of caspase activity carried out using poly(ADP-ribose) polymerase (PARP) as a substrate as well as intracellular caspase assays using a fluorigenic caspase-3 substrate confirmed that caspase-3 is activated in Rat-1 cells undergoing cadmium-induced apoptosis. Both Asp-Glu-Val-Asp-aldehyde (DEVD-cho) and Tyr-Val-Ala-Asp-chloromethylketone (YVAD-cmk), selective inhibitors of caspase-3 and caspase-1, respectively, suppressed significantly cadmium-induced cell death. However, the nonselective caspase inhibitor, z-Val-Ala-Asp-floromethylketone (zVAD-fmk), was the most efficacious agent, almost completely blocking cadmium-induced cell death. Taken together, these results demonstrate that as in other forms of apoptosis, caspases play a central role in cadmium-induced cell death.
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PMID:Cadmium induces caspase-mediated cell death: suppression by Bcl-2. 1077 Nov 29

Peripheral blood mononuclear cells (PBMCs) obtained from patients with advanced melanoma but not from healthy individuals were found to undergo spontaneous ex vivo apoptosis upon incubation in medium. PBMCs were evaluated for evidence of apoptosis using Annexin V binding, caspase-3 activation, and DNA fragmentation (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling). PBMCs of patients with melanoma contained a significantly higher proportion (P = 0.0027) of spontaneously apoptotic cells than PBMCs of controls after 24-h incubation in medium alone. The relative proportion of activated Fas+ and tumor necrosis factor receptor 1-positive (TNFR1+) PBMCs was significantly higher in patients with melanoma than that observed in controls. To demonstrate that the TNF family of receptors and ligands was involved in this type of apoptosis, PBMCs were incubated in the presence of agonistic anti-Fas antibody (CH-11) or TNF-alpha. The proportion of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive PBMCs undergoing spontaneous apoptosis was found to be comparable with that induced by CH-11 antibody or TNF-alpha. Three-color flow cytometry revealed that CD3+ Fas+ T cells were especially sensitive to apoptosis and were preprogrammed in vivo to die. Apoptosis occurred in all subsets of PBMCs but was significantly higher (P = 0.01) in the CD3+ CD8+ T-cell subset in patients relative to controls. In two patients with melanoma, who responded clinically to dendritic cell-based peptide vaccines, the proportion of apoptotic T cells was decreased by half after therapy. In patients who were treated previously with vaccination-based therapies, levels of T-cell apoptosis were lower than in the other melanoma patients. The observed accelerated death of T cells, which are activated and susceptible to apoptosis in patients with melanoma, may contribute to a rapid turnover of immune cells, resulting in a decreased immunocompetence.
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PMID:Spontaneous apoptosis of CD8+ T lymphocytes in peripheral blood of patients with advanced melanoma. 1077 63

Viral protein R (Vpr) of human immunodeficiency virus type 1 inhibits cell proliferation by arresting the cell cycle at the G(2) phase and inducing to apoptosis after G(2) arrest. We have reported previously that C81, a carboxy-terminally truncated form of Vpr, interferes with cell proliferation via a novel pathway that is distinct from G(2) arrest. However, the mechanism of this effect of C81 is unknown. We demonstrate here that C81 can induce apoptosis via G(1) arrest of the cell cycle. Immunostaining for various markers of stages of the cell cycle and flow cytometry analysis of DNA content showed that most HeLa cells that had been transiently transfected with a C81 expression vector were arrested at the G(1) phase and not at the G(2) or S phase of the cell cycle. Staining for annexin V, which binds phosphatidylserine on the plasma membrane, as an early indicator of apoptosis and measurement of the activity of caspase-3, a signaling molecule in apoptotic pathways, indicated that C81 is a strong inducer of apoptosis. Expression of C81 induced the condensation, fragmentation, and clumping of chromatin that are typical of apoptosis. Furthermore, the kinetics of the C81-induced G(1) arrest were closely correlated with changes in the number of annexin V-positive cells and the activity of caspase-3. Replacement of Ile or Leu residues by Pro at positions 60, 67, 74, and 81 within the leucine zipper-like domain of C81 revealed that Ile60, Leu67, and Ile74 play important roles both in the C81-induced G(1) arrest and in apoptosis. Thus, it appears that C81 induces apoptosis through pathways that are identical to those utilized for G(1) arrest of the cell cycle. It has been reported that Ile60, Leu67, and Ile74 also play an important role in the C81-induced suppression of growth. These results suggest that the suppression of growth induced by C81 result in apoptosis that is independent of G(2) arrest of the cell cycle.
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PMID:A carboxy-terminally truncated form of the human immunodeficiency virus type 1 Vpr protein induces apoptosis via G(1) cell cycle arrest. 1084 89

We investigated the in vitro effect of As2O3 on proliferation, cell cycle regulation, and apoptosis in human myeloma cell lines. As2O3 significantly inhibited the proliferation of all of eight myeloma cell lines examined in a dose-dependent manner with IC50 of approximately 1-2 microM. DNA flow cytometric analysis indicated that As2O3 (2 microM) induced a G1 and/or a G2-M phase arrest in these cell lines. To address the mechanism of the antiproliferative effect of As2O3, we examined the effect of As2O3 on cell cycle-related proteins in MC/CAR cells in which both G1 and G2-M phases were arrested. Western blot analysis demonstrated that treatment with As2O3 (2 microM) for 72 h did not change the steady-state levels of CDK2, CDK4, cyclin D1, cyclin E, and cyclin B1 but decreased the levels of CDK6, cdc2, and cyclin A. The mRNA and protein levels of CDKI, p21 were increased by treatment with As2O3, but those of p27 were not. In addition, As2O3 markedly enhanced the binding of p21 with CDK6, cdc2, cyclin E, and cyclin A compared with untreated control cells. Furthermore, the activity of CDK6-associated kinase was reduced in association with hypophosphorylation of Rb protein. The activity of cdc2-associated kinase was decreased, which was accompanied by the up-regulation of cdc2 phosphorylation (cdc2-Tyr15 phosphorylation) resulting from reduction of cdc25B and cdc25C phosphatases. As2O3 also induced apoptosis in MC/CAR cells as evidenced by flow cytometric detection of sub-G1 DNA content and annexin V binding assay. This apoptotic process was associated with down-regulation of Bcl-2, loss of mitochondrial transmembrane potential (delta psi(m)), and an increase of caspase-3 activity. These results suggest that As2O3 inhibits the proliferation of myeloma cells, especially MC/CAR cells, via cell cycle arrest in association with induction of p21 and apoptosis.
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PMID:Arsenic trioxide-mediated growth inhibition in MC/CAR myeloma cells via cell cycle arrest in association with induction of cyclin-dependent kinase inhibitor, p21, and apoptosis. 1085 Apr 58

Hypertonic NaCl upregulated two sensitive and specific biochemical indices of apoptosis, caspase-3 activation and annexin V binding, in a time- and dose-dependent fashion in renal medullary mIMCD3 cells. Pretreatment with urea (200 mM for 30 min) protected from the proapoptotic effect of hypertonic stress (200 mosmol/kgH(2)O) in this model. The protective effect of urea was dose dependent and was effective even when applied a short time (< or =1 h) following NaCl exposure; this protective effect was not observed in the nonrenal 3T3 cell line. In both mIMCD3 and 3T3 cells, urea failed to protect from the proapoptotic stressor, ultraviolet (UV)-B irradiation. The ability of urea to protect from hypertonic stress was approximately comparable to the protective effect of peptide mitogens epidermal growth factor and insulin-like growth factor (IGF), but it potentiated the IGF effect. Interestingly, the tyrosine kinase inhibitor, genistein, potentiated the proapoptotic effect of urea yet abrogated the proapoptotic effect of hypertonic stress. In aggregate, these data indicate that urea protects from the proapoptotic effect of hypertonic stress in a potentially cell type-specific and stimulus-specific fashion.
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PMID:Urea protects from the proapoptotic effect of NaCl in renal medullary cells. 1091 55

The intensity and duration of an inflammatory response depends on the balance of factors that favor perpetuation versus resolution. At sites of inflammation, neutrophils adherent to other cells or matrix components are exposed to tumor necrosis factor-alpha (TNFalpha). Although TNFalpha has been implicated in induction of pro-inflammatory responses, it may also inhibit the intensity of neutrophilic inflammation by promoting apoptosis. Since TNFalpha is not only an important activator of the stress-induced pathways leading to p38 MAPk and c-Jun N-terminal kinase (JNK) but also a potent effector of apoptosis, we investigated the effects of TNFalpha on the JNK pathway in adherent human neutrophils and the potential involvement of this pathway in neutrophil apoptosis. Stimulation with TNFalpha was found to result in beta2 integrin-mediated activation of the cytoplasmic tyrosine kinases Pyk2 and Syk, and activation of a three-part MAPk module composed of MEKK1, MKK7, and/or MKK4 and JNK1. JNK activation was attenuated by blocking antibodies to beta2 integrins, the tyrosine kinase inhibitors, genistein, and tyrphostin A9, a Pyk2-specific inhibitor, and piceatannol, a Syk-specific inhibitor. Exposure of adherent neutrophils to TNFalpha led to the rapid onset of apoptosis that was demonstrated by augmented annexin V binding and caspase-3 cleavage. TNFalpha-induced increases in annexin V binding to neutrophils were attenuated by blocking antibodies to beta2 integrins, and the caspase-3 cleavage was attenuated by tyrphostin A9. Hence, exposure of adherent neutrophils to TNFalpha leads to utilization of the JNK-signaling pathways that may contribute to diverse functional responses including induction of apoptosis and subsequent resolution of the inflammatory response.
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PMID:Tumor necrosis factor-alpha activation of the c-Jun N-terminal kinase pathway in human neutrophils. Integrin involvement in a pathway leading from cytoplasmic tyrosine kinases apoptosis. 1105 15

Multiorgan apoptosis occurs during sepsis. Following cecal ligation and puncture (CLP) in rats, thymocytes underwent apoptosis in a time-dependent manner. C5a blockade dramatically reduced thymocyte apoptosis as measured by thymic weight, binding of annexin V to thymocytes, and laddering of thymocyte DNA. When C5a was generated in vivo by infusion of purified cobra venom factor (CVF), thymocyte apoptosis was significantly increased. Similar results were found when CVF was injected in vivo during the early stages of CLP. In animals 12 hours after induction of CLP, there was an increase in the activities of caspase-3, -6, and -9, but not caspase-1 and -8. Cytosolic cytochrome c levels increased by twofold, whereas mitochondrial levels showed a 50% decrease. Western blot analysis revealed that the content of Bcl-X(L) (but not of Bcl-2, BAX, Bad, and Bim) significantly decreased in thymocytes after CLP. C5a blockade in the sepsis model almost completely inhibited caspase-3, -6, and -9 activation, significantly preserved cytochrome c in the mitochondrial fraction, and restored Bcl-X(L) expression. These data suggest that systemic activation of complement induces C5a-dependent apoptosis of thymocytes and that the blockade of C5a during sepsis rescues thymocytes from apoptosis.
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PMID:Protective effects of anti-C5a in sepsis-induced thymocyte apoptosis. 1108 28

To investigate the mode of zinc-induced cell death, the associated morphological changes, and biological events were examined in zinc-treated Molt-4 cells. Fluorescence microscope observations with double staining of zinc-treated cells with Hoechst 33342 and propidium iodide (PI) indicated that the metal induced both necrosis and apoptosis. To confirm this, cells were stained with both PI and FITC-labeled annexin V, which binds phosphatidylserine, and then analyzed by flow cytometry. The results also confirmed that zinc induces mixed types of cell death, necrosis and apoptosis, and that the former induction occurs earlier and at a greater frequency. Hallmarks of apoptosis such as abnormal chromosome condensation and release of cytochrome c, as well as the appearance of annexin-positive cells, appeared along with the expression of mitochondrial membrane protein 7A6. However, zinc did not induce increases in caspase-3 like protease and caspase-8 activities, and caused slightly hypodiploid cells. Furthermore, the induction of cell death and annexin-positive cells was not blocked by the caspase inhibitors Ac-YVAD-CHO and Ac-DEVD-CHO. These results indicate that zinc induces both necrosis and apoptosis, without caspase-3 activation.
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PMID:Zinc induces mixed types of cell death, necrosis, and apoptosis, in molt-4 cells. 1109 35

Previous studies revealed that expression and activation of cyclooxygenase-2 (Cox-2) conveyed a protective principle in murine macrophages, thus attenuating pro-apoptotic actions of chemotherapeutic agents or programmed cell death as a result of massive nitric oxide (NO) generation. Expression of Cox-2 was achieved by treatment of cells with lipopolysaccharide/interferon-gamma or nontoxic doses of NO releasing agents. We reasoned E-type prostanoid formation, and in turn an intracellular cAMP increase as the underlying protective mechanism. To prove our hypothesis, we analyzed the effects of lipophilic cAMP-analogs on NO, cisplatin, or etoposide induced apoptosis in RAW 264.7 macrophages. Selected apoptotic parameters comprised DNA fragmentation (diphenylamine assay), annexin V staining of phosphatidylserine, caspase activity (quantitated by the cleavage of a fluorogenic caspase-3-like substrate Ac-DEVD-AMC), and mitochondrial membrane depolarisation (delta psi). Western blots detected accumulation of the tumor suppressor protein p53, relocation of cytochrome c to the cytosol, and expression of the anti-apoptotic protein Bcl-xL. Prestimulation with lipophilic cAMP-analogs attenuated apoptosis with the notion that cell death parameters were basically absent. To verify gene induction by cAMP in association with protection we established activation of cAMP response element binding protein (CREB) by gel-shift analysis and moreover, treated macrophages with oligonucleotides containing a cAMP-responsive element (CRE) in order to scavenge CREB. Decoy oligonucleotides, but not control oligonucleotides, attenuated cAMP-evoked protection and reestablished pro-apoptotic parameters. We conclude that gene induction by cAMP protects macrophages towards apoptosis that occurs as a result of excessive NO formation or addition of chemotherapeutica. Attenuating programmed cell death by the cAMP-signaling system may be found in association with Cox-2 expression and tumor formation.
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PMID:Attenuation of macrophage apoptosis by the cAMP-signaling system. 1110 34


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