Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.22.62 (caspase-9)
 7,507 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Shiga-like toxin-producing Escherichia coli causes hemorrhagic colitis and hemolytic-uremic syndrome in association with the production of Shiga-like toxins, which induce cell death via either necrosis or apoptosis. However, the abilities of different Shiga-like toxins to trigger apoptosis and the sequence of intracellular signaling events mediating the death of epithelial cells have not been completely defined. Fluorescent dye staining with acridine orange and ethidium bromide showed that Shiga-like toxin 1 (Stx1) induced apoptosis of HEp-2 cells in a dose- and time-dependent manner. Stx2 also induced apoptosis in a dose-dependent manner. Apoptosis induced by Stx1 (200 ng/ml) and apoptosis induced by Stx2 (200 ng/ml) were maximal following incubation with cells for 24 h (94.3% +/- 1.8% and 81.7% +/- 5.2% of the cells, respectively). Toxin-treated cells showed characteristic features of apoptosis, including membrane blebbing, DNA fragmentation, chromatin condensation, cell shrinkage, and the formation of apoptotic bodies, as assessed by transmission electron microscopy. Stx2c induced apoptosis weakly even at a high dose (1,000 ng/ml for 24 h; 26.7% +/- 1.3% of the cells), whereas Stx2e did not induce apoptosis of HEp-2 cells. Thin-layer chromatography confirmed that HEp-2 cells express the Stx1-Stx2-Stx2c receptor, globotriaosylceramide (Gb3), but not the Stx2e receptor, globotetraosylceramide (Gb4). Western blot analysis of poly(ADP-ribose) polymerase (PARP), a DNA repair enzyme, demonstrated that incubation with Stx1 and Stx2 induced cleavage, whereas incubation with Stx2e did not result in cleavage of PARP. A pan-caspase inhibitor (Z-VAD-FMK) and a caspase-8-specific inhibitor (Z-IETD-FMK) eliminated, in a dose-dependent fashion, the cleavage of PARP induced by Shiga-like toxins. Caspase-8 activation was confirmed by detection of cleavage of this enzyme by immunoblotting. Cleavage of caspase-9 and the proapoptotic member of the Bcl-2 family BID was also induced by Stx1, as determined by immunoblot analyses. We conclude that different Shiga-like toxins induce different degrees of apoptosis that correlates with toxin binding to the glycolipid receptor Gb3 and that caspases play an integral role in the signal transduction cascade leading to toxin-mediated programmed cell death.
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PMID:Escherichia coli shiga-like toxins induce apoptosis and cleavage of poly(ADP-ribose) polymerase via in vitro activation of caspases. 1211 81

Here, we have studied the effects of chemically modified tetracyclines (CMTs) on apoptosis both at the level of the cytoplasmic proteolytic caspase cascade, and on Bcl-2 and c-myc mRNA expression in the J774 macrophage cell line. The results indicate that CMTs induce morphological changes consistent with apoptotic events, as clearly demonstrated both by the acridine orange and ethidium bromide staining, and by TUNEL and fragmentation ELISA assays. Furthermore, the analysis of the cell cycle by flow cytometry shows an evident apoptotic sub-G0G1 peak, without important modifications in the cell cycle distribution. CMTs induce programmed cell death (PCD) in a dose-dependent manner and CMT-8 is the strongest among them. CMT-1 and CMT-8 activate mainly caspase-8 as attested by the inhibitory effects of Z-VAD-fmk and Z-IEDT-fmk on CMT-induced apoptosis. Part of CMT-induced PCD is due to the activation of caspase-9, since it is reduced by the specific caspase-9 inhibitor, Z-LEHD-fmk. Besides, CMTs increase Bcl-2 and c-myc mRNA expression. Collectively, these data indicate that CMTs are potentially anti-tumour agents, since they strongly trigger apoptosis both activating the proteolytic system of the caspase family and modulating genes involved in PCD regulation.
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PMID:Chemically modified tetracyclines induce cytotoxic effects against J774 tumour cell line by activating the apoptotic pathway. 1253 35

Sanguinarine, derived from the root of Sanguinaria canadensis and other poppy fumaria species, possesses strong antimicrobial, anti-inflammatory, and antioxidant properties. We earlier showed that sanguinarine kills human epidermoid carcinoma A431 cells via an induction of apoptosis [N. Ahmad et al., Clin. Cancer Res., 6: 1524-1528, 2000]. In this study, using immortalized human keratinocytes (HaCaT cells), we provide information about mechanism of the antiproliferative effect of sanguinarine. Sanguinarine [0.1 (M-2 (M)] treatment to HaCaT cells was found to inhibit in a dose-dependent manner the cell proliferation and induce apoptosis, as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and ELISA, respectively. Sanguinarine treatment also resulted in a significant cleavage of poly(ADP-ribose) polymerase in HaCaT cells. Because mitochondrial pathway is critical for the regulation of apoptosis, we studied the involvement and regulation of mitochondrial events in sanguinarine-mediated apoptosis of HaCaT cells. As shown by the immunoblot analysis, our data clearly demonstrated that sanguinarine treatment to HaCaT cells resulted in a dose-dependent (a) increase in the level of Bax with a concomitant decrease in Bcl-2 levels and (b) increase in Bax/Bcl-2 ratio. Sanguinarine also resulted in significant increases in the proapoptotic members of Bcl-2 family proteins, i.e., Bak and Bid. This was accompanied by increase in (a) protein expression of cytochrome c and apoptotic protease-activating factor-1 and (b) activity and protein expression of caspase-3, caspase-7, caspase-8, and caspase-9. Taken together, our data showed the involvement of mitochondrial pathway and Bcl-2 family proteins during sanguinarine-mediated apoptosis of immortalized keratinocytes. We suggest that sanguinarine could be developed as a drug for the management of hyperproliferative skin disorders, including skin cancer.
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PMID:Activation of prodeath Bcl-2 family proteins and mitochondrial apoptosis pathway by sanguinarine in immortalized human HaCaT keratinocytes. 1291 70

Normal human mammary epithelial cells (HMECs), unlike estrogen receptor-positive (ER+) breast cancers, typically express low nuclear levels of ER (ER-'poor'). We previously demonstrated that 1.0 microM tamoxifen (Tam) induced apoptosis in ER-'poor' HMECs acutely transduced with human papillomavirus-16 E6 (HMEC-E6) through a rapid mitochondrial signaling pathway. Here, we show that plasma membrane-associated E2-binding sites initiate the rapid apoptotic effects of Tam in HMEC-E6 cells through modulation of AKT activity. At equimolar concentrations, Tam and tamoxifen ethyl bromide (QTam), a membrane impermeant analog of Tam, rapidly induced apoptosis in HMEC-E6 cells associated with an even more rapid decrease in phosphorylation of AKT at serine-473. Treatment of HMEC-E6 cells with 1.0 microM QTam resulted in a 50% decrease in mitochondrial transmembrane potential, sequential activation of caspase-9 and -3, and a 90% decrease in AKT Ser-473 phosphorylation. The effects of both Tam and QTam were blocked by expression of constitutively active AKT (myristoylated AKT or AKT-Thr308Asp/Ser473Asp). These data indicate that Tam and QTam induce apoptosis in HMEC-E6 cells through a plasma membrane-activated AKT-signaling pathway that results in (1) decreased AKT phosphorylation at Ser-473, (2) mitochondrial membrane depolarization, and (3) activated caspase-9 and -3.
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PMID:Tamoxifen and tamoxifen ethyl bromide induce apoptosis in acutely damaged mammary epithelial cells through modulation of AKT activity. 1499 Sep 93

Evodiamine, isolated from a Chinese herbal drug named Wu-Chu-Yu, possesses many biological functions. Recently, it has been reported that Wu-Chu-Yu exerts an antiproliferative effect on several cancers. Prostate carcinoma initially occurs as an androgen-dependent tumor and is the second leading cause of cancer death in American males. In the present study, the effect of evodiamine on the growth of androgen-dependent prostate cancer cell line LNCaP in vitro was examined. Based on [3-(4,5-dimethylthiazol-2-yle)2,5-diphenyltetrazolium bromide] (MTT) assay, evodiamine significantly inhibited the growth of LNCaP cells in a concentration-dependent manner. A significant and concentration-dependent inhibitory effect of evodiamine on LNCaP cell growth was observed at 24 hr and persisted for 96 hr. The examination of lactate dehydrogenase (LDH) assay showed that the cytotoxic effects of evodiamine on LNCaP cells were concentration dependent. Furthermore, we examined the influences of evodiamine on cell death and cell cycle. The flow cytometric analysis of evodiamine-treated cells indicated a block of G2/M phase and an elevated level of DNA fragmentation. The G2/M arrest reached a maximum at 24 hr after evodiamine treatment. The G2/M arrest was accompanied by an elevated p34(cdc2) kinase activity and an increase in the protein expression of cyclin B1 and phosphorylated form of p34(cdc2) (Thr 161). Examination of TUNEL showed that evodiamine-induced apoptosis was observed at 24 hr and extended for 72 hr. Evodiamine elevated caspase-3, and caspase-9 activities and the processing of caspase-3 and caspase-9. These results suggested that evodiamine inhibits the growth of prostate cancer cell line, LNCaP, through an accumulation of cell cycle at G2/M phase and an induction of apoptosis.
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PMID:Inhibitory effects of evodiamine on the growth of human prostate cancer cell line LNCaP. 1514 52

Norcantharidin (NCTD) is the demethylated form of cantharidin, which is the active substance of mylabris. To examine the pathway of NCTD-induced A375-S2 cell death, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-dipheyltetrazolium bromide (MTT) assay, photomicroscopical observation, DNA agarose gel electrophoresis, caspase activity assay and Western blot analysis were carried out. A375-S2 cells treated with NCTD exhibited several typical characteristics of apoptosis. The inhibitory effect of NCTD on human melanoma, A375-S2 cells, was partially reversed by the inhibitors of pan-caspase, caspase-3 and caspase-9. The activities of caspase-3 and -9 were significantly increased after treatment with NCTD at different time. The expression of inhibitor of caspase-activated DNase was decreased in a time-dependent manner, simultaneously, the ratio of Bcl-2/Bax or Bcl-xL/Bax was decreased and the expression ratio of proteins could be reversed by caspase-3 inhibitor. The expression of cytochrome c in cytosol was increased after NCTD treatment and caspase- 3 inhibitor had no significant effect on the up-regulation of cytochrom c. These results suggest that NCTD induced A375-S2 cell apoptosis and the activation of caspase and mitochondrial pathway were involved in the process of NCTD-induced A375-S2 cell apoptosis.
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PMID:Norcantharidin induces human melanoma A375-S2 cell apoptosis through mitochondrial and caspase pathways. 1530 48

We studied the ability of phorbol 12-myristate 13-acetat to prevent erythroid differentiation and apoptosis in erythroleukemic K562 cells induced by cytidine, thymidine, and guanosine. The exposure of cancer cells to combinations of phorbol 12-myrsitate 13-acetate (100 nM) nucleosides for two days led to a loss of hemoglobin production (marker of erythroid differentiation) in cells and increased expression of monocyte-macrophage lineage associated surface antigen CD14. The treatment of K562 cells with nucleosides only was accompanied by the activation of caspase-3 and caspase-9, rather than caspase-6, increased fluorescence of ethidium bromide and DAPI upon binding to DNA, and apoptosis. Intracellular activation of caspase-6, inhibition of caspase-9, a markedly decreased activity of caspase-3 and of fluorescence of DNA-binding dyes, and inhibition of apoptosis were observed when the cells were treated with phorbol 12-myeristet 13-acetate combined with nucleosides.
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PMID:[Phorbol 12-myristate 13-acetate inhibits apoptosis in erythroleukemia K562 cells induced by some nucleosides]. 1580 33

Curcumin, a major yellow pigment and active component of turmeric, has multiple anti-cancer properties. However, its molecular targets and mechanisms of action on human colon adenocarcinoma cells are unknown. In the present study, we examined the effects of curcumin on the proliferation of human colon adenocarcinoma HT-29 cells by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide method and confirmed the curcumin-induced apoptosis by morphology and DNA ladder formation. At the same time, p53, phospho-p53 (Ser15), and other apoptosis-related proteins such as Bax, Bcl-2, Bcl-xL, pro-caspase-3, and pro-caspase-9 were determined by Western blot analysis. The colon adenocarcinoma cells were treated with curcumin (0-75 microM) for 0-24 h. We observed that p53 was highly expressed in HT-29 cells and curcumin could up-regulate the serine phosphorylation of p53 in a time- and concentration-dependent manner. An increase in expression of the pro-apoptotic factor Bax and a decrease in expression of the anti-apoptotic factor Bcl-2 were also observed in a time-dependent manner after exposure of 50 microM curcumin, while the expression of the anti-apoptotic factor Bcl-xL was unchanged. Curcumin could also down-regulate the expression of pro-caspase-3 and pro-caspase-9 in a time-dependent manner. These data suggest a possible underlying molecular mechanism whereby curcumin could induce the apoptosis signaling pathway in human HT-29 colon adenocarcinoma cells by p53 activation and by the regulation of apoptosis-related proteins. This property of curcumin suggests that it could have a possible therapeutic potential in colon adenocarcinoma patients.
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PMID:Curcumin induces human HT-29 colon adenocarcinoma cell apoptosis by activating p53 and regulating apoptosis-related protein expression. 1630 93

In an attempt to dissect the mechanism of Strychnos nux-vomica, a commonly used Chinese folk medicine in the therapy of liver cancer, the cytotoxic effects of four alkaloids in Strychnos nux-vomica, brucine, brucine N-oxide, strychnine, and isostrychnine, on human hepatoma cells (HepG2) were screened by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrasolium bromide (MTT) assay. Brucine, among the four alkaloids, exhibited the strongest toxic effect, the mechanism of which was found to cause HepG2 cell apoptosis, since brucine caused HepG2 cell shrinkage, the formation of apoptotic bodies, DNA fragmentation, cell cycle arrest, as well as phosphatidylserine externalization, all of which are typical characteristics of apoptotic programmed cell death. Brucine-induced HepG2 cell apoptosis was caspase dependent, with caspase-3 activated by caspase-9. Brucine also caused the proteolytic processing of caspase-9. In addition, brucine caused depolarization of the mitochondrial membrane of HepG2 cells, the inhibition of which by cyclosporine A completely abrogated the activation of casapses and release of cytochrome c in brucine-treated HepG2 cells. These findings suggested a pivotal role of mitochondrial membrane depolarization in HepG2 cell apoptosis elicited by brucine. Furthermore, brucine induced a rapid and sustained elevation of intracellular [Ca2+], which compromised the mitochondrial membrane potential and triggered the process of HepG2 cell apoptosis. Finally, Bcl-2 was found to predominately control the whole event of cell apoptosis induced by brucine. The elevation of [Ca2+]i caused by brucine was also suppressed by overexpression of Bcl-2 protein in HepG2 cells. From the facts given above, Ca2+ and Bcl-2 mediated mitochondrial pathway were found to be involved in brucine-induced HepG2 cell apoptosis.
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PMID:The apoptotic effect of brucine from the seed of Strychnos nux-vomica on human hepatoma cells is mediated via Bcl-2 and Ca2+ involved mitochondrial pathway. 1644 26

The present study tests the hypothesis that cerebral hypoxia results in increased ratio of Bax/Bcl-2, activation of caspase-9, lipid peroxidation, and DNA fragmentation in mitochondria of the cerebral cortex of newborn piglets and that the inhibition of nitric oxide synthase by N-nitro-L-arginine during hypoxia will prevent the events leading to mitochondrial DNA fragmentation. To test this hypothesis, six piglets, 3-5 days old, were divided into three groups: normoxic (n=5), hypoxic (n=5), and hypoxic-nitric oxide synthase (n=4). Hypoxic animals were exposed to a FiO2 of 0.6 for 60 min. Nitric oxide synthase (40 mg/kg) was infused over 60 min prior to hypoxia. Tissue hypoxia was confirmed by measuring levels of ATP and phosphocreatine. Cerebral cortical tissue mitochondria were isolated and purified using a discontinuous ficoll gradient. Mitochondrial Bax and Bcl-2 proteins were determined by Western blot. Caspase-9 activity in mitochondria was determined spectro-fluorometrically using fluorogenic substrate for caspase-9. Fluorescent compounds, an index of mitochondrial membrane lipid peroxidation, were determined spectrofluorometrically. Mitochondrial DNA was isolated and separated by electrophoresis on 1% agarose gel and stained with ethidium bromide. ATP levels (micromol/g brain) were 4.52+/-0.34 in normoxic, 1.18+/-0.29 in hypoxic (P<0.05) and 1.00+/-0.26 in hypoxic-nitric oxide synthase animals (P<0.05 vs. normoxic). Phosphocreatine levels (micromol/g brain) were 3.61+/-0.33 in normoxic, 0.70+/-0.20 in hypoxic (P<0.05 vs. normoxic) and 0.57+/-0.14 in hypoxic-nitric oxide synthase animals (P<0.05 vs. normoxic, P=NS vs. hypoxic). Bax density in mitochondrial membranes was 160+/-28 in normoxic and 324+/-65 in hypoxic (P<0.001 vs. normoxic). Bcl-2 density mitochondria was 96+/-18 in normoxic and 98+/-20 in hypoxic (P=NS vs. normoxic). Mitochondrial caspase-9 activity (nmol/mg protein/h) was 1.32+/-0.23 in normoxic and 2.25+/-0.24 in hypoxic (P<0.01 vs. normoxic). Levels of fluorescent compounds (microg of quinine sulfate/g protein) were 12.48+/-4.13 in normoxic and 37.92+/-7.62 in hypoxic (P=0.003 vs. normoxic). Densities (ODxmm2) of low molecular weight DNA fragments were 143+/-38 in normoxic, 365+/-152 in hypoxic, (P<0.05 vs. normoxic) and 163+/-25 in hypoxic-nitric oxide synthase animals (P<0.05 vs. hypoxic, P=NS vs. normoxic). The data demonstrate that hypoxia results in increased mitochondrial proapoptotic protein Bax, increased mitochondrial caspase-9 activity, increased mitochondrial lipid peroxidation, and increased fragmentation of DNA in mitochondria of the cerebral cortex of newborn piglets. The administration of a nitric oxide synthase inhibitor, nitric oxide synthase, prior to hypoxia prevented fragmentation of mitochondrial DNA, indicating that the hypoxia-induced mitochondrial DNA fragmentation is NO-mediated. We propose that NO free radicals generated during hypoxia lead to NO-mediated altered expression of Bax leading to increased ratio of pro-apoptotic/anti-apoptotic protein resulting in modification of mitochondrial membrane, and subsequently Ca2+-influx and fragmentation of mitochondrial DNA.
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PMID:Hypoxia-induced Bax and Bcl-2 protein expression, caspase-9 activation, DNA fragmentation, and lipid peroxidation in mitochondria of the cerebral cortex of newborn piglets: the role of nitric oxide. 1677 44


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