EC:3.4.22.56 - FACTA Search

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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 present study examined whether X-ray- and CDDP-sensitivities depend on p53 gene status in human squamous cell carcinoma of the head and neck (SAS cells) showing dominant negative nature of mutant p53 protein. SAS cells were transfected with a vector carrying a mutant p53 gene (SAS/Trp248 cells) or neomycin resistant gene control vector (SAS/neo cells). Sensitivities of the transfected cells to X-ray or CDDP were measured with colony formation assay. The incidence of apoptosis by X-ray or CDDP was analyzed with Hoechst staining or DNA ladder formation assay. The activation of caspase-3 was estimated as an indicator of apoptosis by the detection of fragmentation of caspase-3 or poly (ADP ribose) polymerase (PARP) with Western blot. SAS/Trp248 cells showed X-ray- and CDDP-resistance due to the dominant negative nature of mutant p53, compared with SAS/neo cells. The incidence of DNA ladders and apoptotic bodies increased markedly in SAS/neo cells after X-ray irradiation or CDDP treatment, but increased only slightly in SAS/Trp248 cells. Fragmentation of caspase-3 and PARP was observed in SAS/neo cells, but almost no such fragmentation was observed in SAS/Trp248 cells after X-ray irradiation or CDDP treatment. The present results strongly suggest that the X-ray- and CDDP-sensitivities of human squamous cell carcinomas are p53-dependent, and that the sensitivities are tightly correlated with the induction of apoptosis through caspase-3 activation. The p53-dependent X-ray- or CDDP-sensitivity was supported by results from p53-null human lung cancer H1299 cells which were transfected with wild-type or mutant p53 gene.
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PMID:Transfection of mutant p53 gene depresses X-ray- or CDDP-induced apoptosis in a human squamous cell carcinoma of the head and neck. 1210 96

Monocytes/macrophages are thought to be involved in Acanthamoeba infections. The aim of this work was to study whether soluble metabolites (ADP and other compounds) released by Acanthamoeba castellanii trophozoites could induce morphological and biochemical changes in human monocytic cells in vitro. We demonstrate here that ADP constitutively released in the medium by A. castellanii, interacting with specific P2y(2) purinoceptors expressed on the monocytic cell membrane, caused a biphasic rise in [Ca(2+)](i), morphological changes characteristics of cells undergoing apoptosis, caspase-3 activation, and secretion of tumor necrosis factor alpha (TNF-alpha). The same results were found in monocytes exposed to purified ADP. Cell damage and TNF-alpha release induced by amoebic ADP were blocked by the P2y(2) inhibitor suramin. Other metabolites contained in amoebic cell-free supernatants, with molecular masses of, respectively, >30 kDa and between 30 and 10 kDa, also caused morphological modifications and activation of intracellular caspase-3, characteristics of programmed cell death. Nevertheless, mechanisms by which these molecules trigger cell damage appeared to differ from that of ADP. In addition, other amoebic thermolable metabolites with molecular masses of <10 kDa caused the secretion of interleukin-1beta. These findings suggest that pathogenic free-living A. castellanii by release of ADP and other metabolites lead to human monocytic cell death through apoptosis and stimulate the secretion of proinflammatory cytokines.
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PMID:ADP and other metabolites released from Acanthamoeba castellanii lead to human monocytic cell death through apoptosis and stimulate the secretion of proinflammatory cytokines. 1211 53

Pierisin-1, a 98-kDa protein that induces apoptosis in mammalian cell lines, is capable of being incorporated into cells where it ADP-ribosylates guanine residues in DNA. To investigate the apoptotic pathway induced by this unique protein, the bcl-2 gene was transfected into HeLa cells. Cy2-fluorescent pierisin-1 was incorporated into the resultant cells expressing Bcl-2 protein and ADP-ribosylated dG was detected to almost the same extent as in parent cells. However, bcl-2-transfected HeLa cells did not display apoptotic morphological changes, PARP cleavage, and DNA fragmentation, indicating acquisition of resistance. In parent HeLa cells, activation of caspase-9 and release of cytochrome c were observed after 8h treatment with 0.5ng/ml pierisin-1. Caspase substrate assays revealed further cleavage of Ac-DEVD-pNA, Ac-VDVAD-pNA, and Ac-VEID-pNA, suggesting activation of caspase-2, -3, and -6 in pierisin-1-treated HeLa cells. The caspase-3 inhibitor, Ac-DEVD-CHO, was also found to inhibit apoptosis. In contrast, this caspase activation was not observed in bcl-2-transfected HeLa cells. Our results thus indicate that pierisin-1-induced apoptosis is mediated primarily via a mitochondrial pathway involving Bcl-2 and caspases.
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PMID:Bcl-2 blocks apoptosis caused by pierisin-1, a guanine-specific ADP-ribosylating toxin from the cabbage butterfly. 1214 21

We tested the effects of hydroxychloroquine (HCQ), an anti-rheumatic drug, on the viability of chronic lymphocytic leukemia (CLL) cells. HCQ induced a decrease in cell viability in a dose- and time-dependent manner. The mean LC50 calculated for the cells of 20 patients was 32 +/- 7 microg/ml (range, 10-75 microg/ml). We observed a large increase in apoptotic cell number after 24 h of incubation with 50 microg/ml HCQ (55 +/- 6 vs. 23 +/- 3% in medium alone, p < 0.001). Indeed, HCQ in leukemic cells induced the features of apoptosis (cell shrinkage, decrease in mitochondrial transmembrane potential, phosphatidylserine externalization, chromatin condensation and DNA fragmentation). HCQ had marked selective cytotoxicity when compared with normal blood mononuclear cells, in which the LC50 was >100 microg/ml at 24 h. HCQ induced the proteolytic cleavage of poly(ADP(adenosine 5'-diphosphate)ribose) polymerase (PARP) and increased the activity of caspase-3. The expression of bcl-2 and bax proteins was significantly modified after incubation with the drug and HCQ activity against CLL cells occurred independently of the presence of IL-4, sCD40L and bone marrow stromal cells.
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PMID:Hydroxychloroquine-induced apoptosis of chronic lymphocytic leukemia involves activation of caspase-3 and modulation of Bcl-2/bax/ratio. 1214 91

The cytotoxic effect of the chemotherapeutic drug etoposide (VP-16) is thought to result from its ability to induce DNA damage and thereby to trigger apoptosis. Internucleosomal DNA fragmentation occurs late during apoptosis in many cell types. However, whereas human osteosarcoma cells undergo internucleosomal DNA fragmentation during staurosporine-induced apoptosis, they fail to do so in response to VP-16. Recently, we showed that these cells also do not express the poly(ADP-ribosyl)ation-regulated Ca(2+)- and Mg(2+)-dependent endonuclease DNAS1L3. The possibility that this deficiency underlies the failure of these cells to undergo internucleosomal DNA fragmentation in response to VP-16 was investigated. The proteolytic processing and consequent activation of procaspase-3, cleavage of the inhibitory subunit of DNA fragmentation factor, and the degradation of DNA into 50-kb fragments occurred similarly in osteosarcoma cells exposed to either staurosporine or VP-16. However, the additional processing of the 50-kb DNA fragments to oligonucleosomal fragments was not apparent in the VP-16-treated cells. Ectopic expression of DNAS1L3 conferred on osteosarcoma cells the ability to undergo VP-16-induced internucleosomal DNA fragmentation. Furthermore, expression of DNAS1L3 markedly potentiated the cytotoxic effect of VP-16 in these cells. Both DNAS1L3-mediated and staurosporine-induced internucleosomal DNA fragmentation were Ca(2+) dependent, but only the DNAS1L3-mediated DNA cleavage was blocked by expression of a caspase-3-resistant mutant of poly(ADP-ribose) polymerase-1. The present work results suggest a direct relation between the activity of a chemotherapeutic drug (VP-16) and a specific endonuclease (DNAS1L3). They also indicate that internucleosomal DNA fragmentation plays an active role in apoptosis and that the failure of cancer cells to undergo such DNA degradation may contribute to the development of resistance to chemotherapeutic drugs.
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PMID:The Poly(ADP-ribose) polymerase-1-regulated endonuclease DNAS1L3 is required for etoposide-induced internucleosomal DNA fragmentation and increases etoposide cytotoxicity in transfected osteosarcoma cells. 1215 52

This study was designed to evaluate the synergistic cytotoxicity of herb prescription, Palgin, in adriamycin-treated cancer cells. The combination of Palgin and adriamycin synergistically augmented the cytotoxicity of Chang and HL-60 cells, but not in Hep3B and Alexander cells. The cytotoxicity of two drugs was revealed as apoptosis characterized by nuclear fragmentation. The apoptotic cell death was accompanied by the activation of caspase-3 as well as cleavage of poly(ADP) ribose polymerase (PARP) in Chang cells. Interestingly, a synergistic increase in apoptosis by the combination of two drugs was accompanied by the enhancement of Fas and Fas ligand (FasL) expression in Chang cells. Taken together, the combination of Palgin and adriamycin significantly augmented the apoptotic cytotoxicity of Fas-positive cells, such as Chang and HL-60 cells, via activation of caspase signaling pathway. This notion will provide a new trial to treat cancer patients in clinical fields as a complementary treatment of Western and Oriental medicine.
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PMID:Palgin sensitizes the adriamycin-induced apoptosis via the enhancement of Fas/Fas ligand expression. 1223

This report is focused on the apoptotic effect induced by MG132, an inhibitor of 26S proteasome, in human hepatoma HepG2 cells. The results were compared with those obtained with non-transformed human Chang liver cells. MG132 reduced the viability of HepG2 cells in a time- and dose-dependent manner. The effect was in tight connection with the induction of apoptosis, as indicated by fluorescence microscopy and cytometric analysis, and was accompanied by a remarkable increase in the production of H2O2 and a reduction in mitochondrial transmembrane potential (Deltapsim). In addition cell death was prevented by antioxidants such as GSH, N-acetylcysteine or catalase. Western blot analysis showed that HepG2 cells contain a very low level of Bcl-2 and a much higher level of Bcl-XL, another antiapoptotic factor of the same family. When the cells were exposed to MG132 the level of Bcl-XL diminished, while a new band, corresponding to the expression of the proapoptotic protein Bcl-XS was detected. MG132 also caused the release of cytochrome c from mitochondria and the activation of caspase-3 with the consequent degradation of poly-ADP ribose polymerase (PARP). The observation that the broad spectrum caspase inhibitor z-VAD markedly reduced the apoptotic effect of the drug clearly demonstrated that caspases play an important role in MG132-induced apoptosis. MG132 exerted a modest effect on the viability of Chang liver cells which primarily depended on the G2/M arrest of cell cycle while only a small percentage of apoptotic cells was found. The remarkable differences in the effects induced by MG132 in Chang liver cells and HepG2 cells made us hypothesise the potential use of proteasome inhibitors in hepatocarcinoma therapy.
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PMID:Apoptosis induced in hepatoblastoma HepG2 cells by the proteasome inhibitor MG132 is associated with hydrogen peroxide production, expression of Bcl-XS and activation of caspase-3. 1223 27

Apoptosis of retinal endothelial cells and pericytes is postulated to contribute to the development of retinopathy in diabetes. The goal of this study is to investigate diabetes-induced activation of retinal caspase-3, an apoptosis executer enzyme, in retina, and examine the effects of antioxidants on the activation. Caspase-3 activation was determined in the retina of alloxan diabetic rats (2-14 months duration) and in the isolated retinal capillary cells (endothelial cells and pericytes) by measuring cleavage of caspase-3 specific fluorescent substrate, and cleavage of caspase-3 holoenzyme and poly (ADP ribosyl) polymerase. Effect of antioxidants on the activation of caspase-3 was determined by feeding a group of diabetic rats diet supplemented with a comprehensive mixture of antioxidants, including Trolox, alpha-tocopherol, N-acetyl cysteine, ascorbic acid, beta-carotene and selenium for 2-14 months, and also under in vitro conditions by incubating isolated retinal capillary cells with antioxidants with wide range of actions. Caspase-3 was activated in the rat retina at 14 months of diabetes (P < 0.05 vs. normal), but not at 2 months of diabetes, and administration of antioxidants for the entire duration inhibited this activation. In the isolated retinal capillary cells incubated in 25 mM glucose medium, caspase-3 activity was increased by 50% compared to the cells incubated in 5 mM glucose (P < 0.02), and antioxidants or caspase-3 inhibitor inhibited this increase. Our results suggest that increased oxidative stress in diabetes is involved in the activation of retinal caspase-3 and apoptosis of endothelial cells and pericytes. Antioxidants might be inhibiting the development of diabetic retinopathy by inhibiting microvascular apoptosis.
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PMID:Diabetes-induced activation of caspase-3 in retina: effect of antioxidant therapy. 1244 25

It is well known that diabetes aggravates brain damage in experimental and clinical stroke subjects. Diabetes accelerates maturation of neuronal damage, increases infarct volume, and induces postischemic seizures. The mechanism by which diabetes increases ischemic brain damage is still elusive. Our previous experiments indicate that mitochondria dysfunction may play a role in neuronal death. The objective of this study is to determine whether streptozotocin-induced diabetes activates cell death pathway after a brief period of focal cerebral ischemia. Both diabetic and nondiabetic rats were subjected to 30 min of transient middle cerebral artery occlusion, followed by 0, 0.5, 3, and 6 h of reperfusion. We first determined the pathological outcomes after 7 days of recovery by histopathology, and then detected key components of programmed cell death pathway using immunocytochemistry coupled with confocal laser-scanning microscopy and Western blot analysis. The results show that the cytosolic cytochrome c increased mildly after reperfusion in nondiabetic samples. This increase was markedly enhanced in diabetic rats in both ischemic focus and penumbra. Subsequently, caspase-3 was activated and poly-ADP ribose polymerase (PARP) was cleaved. Our results suggest that activation of apoptotic cell death pathway may play a pivotal role in exaggerating brain damage in diabetic subjects.
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PMID:Diabetes activates cell death pathway after transient focal cerebral ischemia. 1254 Jun 24

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been shown to exert potent cytotoxic activity against many tumor cells but not normal cells. However, some tumor cells are resistant to TRAIL, and it has not been determined how this occurs. In the present study, we obtained three subgroups of Jurkat clones with TRAIL-sensitive, -partial resistant and -resistant phenotypes. We found that most TRAIL-resistant and -partial resistant clones expressed low levels of DR5, whereas most TRAIL-sensitive clones expressed high levels of Death Receptor (DR5). However, there were clones with a range of different TRAIL-sensitivities that had similar levels of DR5 expression. The expression levels of DR4 and the decoy receptors, DcR1 and DcR2, did not correlate with TRAIL sensitivities. We also compared the subgroups in terms of the expression of Fas-associated death domain protein (FADD), the levels of activation of Receptor Interacting Protein (RIP) and caspases, and cleavage of Poly (ADP-Ribose)Polymerase (PARP). Basal expression levels of FADD were not significantly different among the subgroups. After treatment with TRAIL, both TRAIL-sensitive and partial resistant clones showed high levels of activation of caspase-3, caspase-8, RIP and PARP. Relative basal level and induced level of Phosphoprotein over Expressed in Diabetes/Phosphoprotein Enriched in Astrocytes (PED/PEA-15) after TRAIL treatment were compared in the clones. Basal levels of PED/PEA-15 expression were similar among sensitive, partial resistant and resistant clones. TRAIL did not change the PED/PEA-15 level in the clones. In addition, transduction and expression of the dominant negative form of the I-kBalpha gene did not change TRAIL-sensitivities. Our results showed that the expression levels of DR5, the activation levels of caspase-8, -3 and RIP were critical factors in determining TRAIL-sensitivities in Jurkat cells. The results of our study also suggest that cells with different TRAIL-sensitivities arise through multiple mechanisms even within a single cell line.
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PMID:Analysis of the phenotypes of Jurkat clones with different TRAIL-sensitivities. 1270 64

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