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Sample records for caspase-8 induce terminal

  1. A novel caspase 8 selective small molecule potentiates TRAIL-induced cell death.

    Science.gov (United States)

    Bucur, Octavian; Gaidos, Gabriel; Yatawara, Achani; Pennarun, Bodvael; Rupasinghe, Chamila; Roux, Jérémie; Andrei, Stefan; Guo, Bingqian; Panaitiu, Alexandra; Pellegrini, Maria; Mierke, Dale F; Khosravi-Far, Roya

    2015-05-11

    Recombinant soluble TRAIL and agonistic antibodies against TRAIL receptors (DR4 and DR5) are currently being created for clinical cancer therapy, due to their selective killing of cancer cells and high safety characteristics. However, resistance to TRAIL and other targeted therapies is an important issue facing current cancer research field. An attractive strategy to sensitize resistant malignancies to TRAIL-induced cell death is the design of small molecules that target and promote caspase 8 activation. For the first time, we describe the discovery and characterization of a small molecule that directly binds caspase 8 and enhances its activation when combined with TRAIL, but not alone. The molecule was identified through an in silico chemical screen for compounds with affinity for the caspase 8 homodimer's interface. The compound was experimentally validated to directly bind caspase 8, and to promote caspase 8 activation and cell death in single living cells or population of cells, upon TRAIL stimulation. Our approach is a proof-of-concept strategy leading to the discovery of a novel small molecule that not only stimulates TRAIL-induced apoptosis in cancer cells, but may also provide insights into the structure-function relationship of caspase 8 homodimers as putative targets in cancer.

  2. Salmonella infection induces recruitment of Caspase-8 to the inflammasome to modulate IL-1β production.

    Science.gov (United States)

    Man, Si Ming; Tourlomousis, Panagiotis; Hopkins, Lee; Monie, Tom P; Fitzgerald, Katherine A; Bryant, Clare E

    2013-11-15

    Nucleotide-binding oligomerization domain-like receptors (NLRs) detect pathogens and danger-associated signals within the cell. Salmonella enterica serovar Typhimurium, an intracellular pathogen, activates caspase-1 required for the processing of the proinflammatory cytokines, pro-IL-1β and pro-IL-18, and pyroptosis. In this study, we show that Salmonella infection induces the formation of an apoptosis-associated specklike protein containing a CARD (ASC)-Caspase-8-Caspase-1 inflammasome in macrophages. Caspase-8 and caspase-1 are recruited to the ASC focus independently of one other. Salmonella infection initiates caspase-8 proteolysis in a manner dependent on NLRC4 and ASC, but not NLRP3, caspase-1 or caspase-11. Caspase-8 primarily mediates the synthesis of pro-IL-1β, but is dispensable for Salmonella-induced cell death. Overall, our findings highlight that the ASC inflammasome can recruit different members of the caspase family to induce distinct effector functions in response to Salmonella infection.

  3. The equine arteritis virus induces apoptosis via caspase-8 and mitochondria-dependent caspase-9 activation.

    Science.gov (United States)

    St-Louis, Marie-Claude; Archambault, Denis

    2007-10-10

    We have previously showed that equine arteritis virus (EAV), an arterivirus, induces apoptosis in vitro. To determine the caspase activation pathways involved in EAV-induced apoptosis, target cells were treated with peptide inhibitors of apoptosis Z-VAD-FMK (pan-caspase inhibitor), Z-IETD-FMK (caspase-8-specific inhibitor) or Z-LEHD-FMK (caspase-9-specific inhibitor) 4 h prior to infection with the EAV T1329 Canadian isolate. Significant inhibition of apoptosis was obtained with all peptide inhibitors used. Furthermore, apoptosis was inhibited in cells expressing the R1 subunit of herpes simplex virus type 2 ribonucleotide reductase (HSV2-R1) or hsp70, two proteins which are known to inhibit apoptosis associated with caspase-8 activation and cytochrome c release-dependent caspase-9 activation, respectively. Given the activation of Bid and the translocation of cytochrome c within the cytoplasm, the overall results indicate that EAV induces apoptosis initiated by caspase-8 activation and subsequent mitochondria-dependent caspase-9 activation.

  4. Human cell-death-inducing DFF45-1ike effector C induces apoptosis via caspase-8

    Institute of Scientific and Technical Information of China (English)

    Xin Tang; Zhen Xing; Hong Tang; Liang Liang; Mujun Zhao

    2011-01-01

    Human cell-death-inducing DNA-fragmentation-factor (DFF45)-like effector C (CIDEC) is a potent apoptotic inducer.Previous studies have indicated that the Fatspecific protein 27 (Fsp27),a mouse homolog of CIDEC,induces apoptosis via caspase-3,-7,and -9 and triggers the release of cytocbrome c from mitochondria,which implies that the mitochondrial pathway is involved in Fsp27-induced apoptosis,in the current study,we found that CIDEC-inducedapoptosiswasmediatedby caspase-8.The caspase inhibitor assay showed that CIDEC-induced apoptosis was dramatically reduced in the presence of the general caspase inhibitor,the caspase-3 inhibitor,and the caspase-8 inhibitor,whereas the caspase-9 inhibitor only weakly inhibited CIDEC-induced apoptosis.These results confirmed that the activation of caspase-3 and caspase-8 were involved in CIDEC-induced apoptosis.Moreover,in caspase-3- or caspase-8-deficient cells,CIDEC-induced apoptosis were dramatically decreased,which demonstrated that CIDEC-induced apoptosis might require the activation of caspase-3 and caspase-8.Because caspase-8 in general is a key effecter of death-receptor pathway and activated by Fas-Associated protein with Death Domain (FADD),we examined whether FADD was involved in CIDEC-induced apoptosis.Our results demonstrated that CIDEC-induced apoptosis was independent of FADD,suggesting that CIDEC-induced apoptosis might be in a death-receptor-independent,caspase-8-dependent manner.It was also found that the region of amino acid 168-200 in carboxyl domain of CIDEC was critical for its crucial pro-apoptotic function.

  5. Caspase-8 regulation by direct interaction with TRAF6 in T cell receptor-induced NF-kappaB activation.

    Science.gov (United States)

    Bidère, Nicolas; Snow, Andrew L; Sakai, Keiko; Zheng, Lixin; Lenardo, Michael J

    2006-08-22

    Triggering of lymphocyte antigen receptors is the critical first step in the adaptive immune response against pathogens. T cell receptor (TCR) ligation assembles a large membrane signalosome, culminating in NF-kappaB activation [1,2]. Recently, caspase-8 was found to play a surprisingly prominent role in lymphocyte activation in addition to its well-known role in apoptosis [3]. Caspase-8 is activated after TCR stimulation and nucleates a complex with B cell lymphoma 10 (BCL10), paracaspase MALT1, and the inhibitors of kappaB kinase (IKK) complex [4]. We now report that the ubiquitin ligase TRAF6 binds to active caspase-8 upon TCR stimulation and facilitates its movement into lipid rafts. We identified in silico two putative TRAF6 binding motifs in the caspase-8 sequence and found that mutation of critical residues within these sites abolished TRAF6 binding and diminished TCR-induced NF-kappaB activation. Moreover, RNAi-mediated silencing of TRAF6 abrogated caspase-8 recruitment to the lipid rafts. Protein kinase Ctheta (PKCtheta), CARMA1, and BCL10 are also required for TCR-induced caspase-8 relocation, but only PKCtheta and BCL10 control caspase-8 activation. Our results suggest that PKCtheta independently controls CARMA1 phosphorylation and BCL10-dependent caspase-8 activation and unveil an essential role for TRAF6 as a critical adaptor linking these two convergent signaling events.

  6. Occurrence of pre-MBT synthesis of caspase-8 mRNA and activation of caspase-8 prior to execution of SAMDC (S-adenosylmethionine decarboxylase)-induced, but not p53-induced, apoptosis in Xenopus late blastulae.

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    Shiokawa, Koichiro; Takayama, Eiji; Higo, Takayasu; Kuroyanagi, Shinsaku; Kaito, Chikara; Hara, Hiroshi; Kajitani, Masayuki; Sekimizu, Kazuhisa; Tadakuma, Takushi; Miura, Kin-Ichiro; Igarashi, Kazuei; Yaoita, Yoshio

    2005-10-21

    Overexpression of S-adenosylmethionine decarboxylase (SAMDC) in Xenopus fertilized eggs activates caspase-9 and executes maternal program of apoptosis shortly after midblastula transition (MBT). We find that overexpression of caspase-8 and p53, like that of SAMDC, induces apoptosis in Xenopus late blastulae. The apoptosis induced by p53 was abolished by injection of mRNA for xdm-2, a negative regulator of p53, and by injection of a peptide inhibitor or a dominant-negative type mutant of caspase-9, but not caspase-8. The apoptosis induced by SAMDC was not abolished by injection of xdm-2 mRNA, but was abolished by injection of a peptide inhibitor or a dominant-negative type mutant mRNA of both caspase-9 and caspase-8. Unlike caspase-9 mRNA, caspase-8 mRNA did not occur as a maternal mRNA rather induced to be expressed during cleavage stage (pre-MBT stage) by overexpression of SAMDC but not p53. Furthermore, while activities to process procaspase-8 and procaspase-9 appeared in SAMDC-overexpressed apoptotic embryos, the activity to process procaspase-8 did not appear in p53-overexpressed apoptotic embryos. We conclude there are at least two pathways in the execution of the maternal program of apoptosis in Xenopus embryos; one being through do novo expression of caspase-8 gene during cleavage stage, and the other without involvement of caspase-8.

  7. Amplification activation loop between caspase-8 and -9 dominates artemisinin-induced apoptosis of ASTC-a-1 cells.

    Science.gov (United States)

    Xiao, Fenglian; Gao, Weijie; Wang, Xiaoping; Chen, Tongsheng

    2012-06-01

    Although caspases have been demonstrated to be involved in artemisinin (ARTE)-induced apoptosis, their exact functions are not well understood. The aim of this report is to explore the roles of caspase-8, -9 and -3 during ARTE-induced apoptosis in human lung adenocarcinoma (ASTC-a-1) cells. ARTE treatment induces a rapid generation of reactive oxygen species (ROS), and ROS-dependent apoptosis as well as the activation of caspase-8, -9 and -3 via time- and dose-dependent fashion. Of upmost importance, inhibition of caspase-8 or -9, but not caspase-3, almost completely blocks the ARTE-induced not only activation of the caspase-8, -9 and -3 but also apoptosis. In addition, the apoptotic process triggered by ARTE does not involve the Bid cleavage, tBid translocation, significant loss of mitochondrial membrane potential and cytochrome c release from mitochondria. Moreover, silencing Bax/Bak does not prevent the ATRE-induced cell death as well as the activation of caspase-8, -9 and -3. Collectively, our data firstly demonstrate that ARTE triggers a ROS-mediated positive feedback amplification activation loop between caspase-8 and -9 independent of mitochondria, which dominantly mediated the ARTE-induced apoptosis via a caspase-3-independent apoptotic pathway in ASTC-a-1 cells. Our findings imply a potential to develop new derivatives from artemisinin to effectively initiate the amplification activation loop of caspases.

  8. Proapoptotic chemotherapeutic drugs induce noncanonical processing and release of IL-1β via caspase-8 in dendritic cells.

    Science.gov (United States)

    Antonopoulos, Christina; El Sanadi, Caroline; Kaiser, William J; Mocarski, Edward S; Dubyak, George R

    2013-11-01

    The identification of noncanonical (caspase-1-independent) pathways for IL-1β production has unveiled an intricate interplay between inflammatory and death-inducing signaling platforms. We found a heretofore unappreciated role for caspase-8 as a major pathway for IL-1β processing and release in murine bone marrow-derived dendritic cells (BMDC) costimulated with TLR4 agonists and proapoptotic chemotherapeutic agents such as doxorubicin (Dox) or staurosporine (STS). The ability of Dox to stimulate release of mature (17-kDa) IL-1β was nearly equivalent in wild-type (WT) BMDC, Casp1(-/-)Casp11(-/-) BMDC, WT BMDC treated with the caspase-1 inhibitor YVAD, and BMDC lacking the inflammasome regulators ASC, NLRP3, or NLRC4. Notably, Dox-induced production of mature IL-1β was temporally correlated with caspase-8 activation in WT cells and greatly suppressed in Casp8(-/-)Rip3(-/-) or Trif(-/-) BMDC, as well as in WT BMDC treated with the caspase-8 inhibitor, IETD. Similarly, STS stimulated robust IL-1β processing and release in Casp1(-/-)Casp11(-/-) BMDC that was IETD sensitive. These data suggest that TLR4 induces assembly of caspase-8-based signaling complexes that become licensed as IL-1β-converting enzymes in response to Dox and STS. The responses were temporally correlated with downregulation of cellular inhibitor of apoptosis protein 1, suggesting suppressive roles for this and likely other inhibitor of apoptosis proteins on the stability and/or proteolytic activity of the caspase-8 platforms. Thus, proapoptotic chemotherapeutic agents stimulate the caspase-8-mediated processing and release of IL-1β, implicating direct effects of such drugs on a noncanonical inflammatory cascade that may modulate immune responses in tumor microenvironments.

  9. Interaction between caspase-8 activation and endoplasmic reticulum stress in glycochenodeoxycholic acid-induced apoptotic HepG2 cells.

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    Iizaka, Toru; Tsuji, Mayumi; Oyamada, Hideto; Morio, Yuri; Oguchi, Katsuji

    2007-11-30

    The accumulation of hydrophobic bile acid, such as glycochenodeoxycholic acid (GCDCA), in the liver has been thought to induce hepatocellular damage in human chronic cholestatic liver diseases. We previously reported that GCDCA-induced apoptosis was promoted by both mitochondria-mediated and endoplasmic reticulum (ER) stress-associated pathways in rat hepatocytes. In this study, we elucidated the relationship between these pathways in GCDCA-induced apoptotic HepG2 cells. HepG2 cells were treated with GCDCA (100-500microM) with or without a caspase-8 inhibitor, Z-IETD-fluoromethyl ketone (Z-IETD-FMK) (30microM) for 3-24h. We demonstrated the presence of both apoptotic pathways in these cells; that is, we showed increases in cleaved caspase-3 proteins, the release of cytochrome c from mitochondria, and the expression of ER resident molecular chaperone Bip mRNA and ER stress response-associated transcription factor Chop mRNA. On the other hand, pretreatment with Z-IETD-FMK significantly reduced the increases, compared with treatment with GCDCA alone. Immunofluorescence microscopic analysis showed that treatment with GCDCA increased the cleavage of BAP31, an integral membrane protein of ER, and pretreatment with Z-IETD-FMK suppressed the increase of caspase-8 and BAP31 cleavage. In conclusion, these results suggest that intact activated caspase-8 may promote and amplify the ER stress response by cleaving BAP31 in GCDCA-induced apoptotic cells.

  10. Diatom-derived oxylipins induce cell death in sea urchin embryos activating caspase-8 and caspase 3/7

    OpenAIRE

    N. Ruocco; Varella, S; Romano, G.; Ianora, A.; Bentley, M. G.; Somma, D.; Leonardi, A.; Mellone, S.; Zuppa, A; Costantini, M

    2016-01-01

    Diatoms are an important class of unicellular algae that produce bioactive secondary metabolites withcytotoxic activity collectively termed oxylipins, including polyunsaturated aldehydes (PUAs), hydroxy-acids (HEPEs), oxo-acids and epoxyalcohols. Previous results showed that at higher concentrations, thePUA decadienal induced apoptosis on copepods and sea urchin embryos via caspase-3 activation; atlower concentrations decadienal affected the expression levels of the caspase-8 gene in embryos ...

  11. Diatom-derived oxylipins induce cell death in sea urchin embryos activating caspase-8 and caspase 3/7.

    Science.gov (United States)

    Ruocco, Nadia; Varrella, Stefano; Romano, Giovanna; Ianora, Adrianna; Bentley, Matt G; Somma, Domenico; Leonardi, Antonio; Mellone, Stefano; Zuppa, Antonio; Costantini, Maria

    2016-07-01

    Diatoms are an important class of unicellular algae that produce bioactive secondary metabolites with cytotoxic activity collectively termed oxylipins, including polyunsaturated aldehydes (PUAs), hydroxyacids (HEPEs), oxo-acids and epoxyalcohols. Previous results showed that at higher concentrations, the PUA decadienal induced apoptosis on copepods and sea urchin embryos via caspase-3 activation; at lower concentrations decadienal affected the expression levels of the caspase-8 gene in embryos of the sea urchin Paracentrotus lividus. In the present work, we studied the effects of other common oxylipins produced by diatoms: two PUAs (heptadienal and octadienal) and four hydroxyacids (5-, 9- 11- and 15-HEPE) on P. lividus cell death and caspase activities. Our results showed that (i) at higher concentrations PUAs and HEPEs induced apoptosis in sea urchin embryos, detected by microscopic observation and through the activation of caspase-3/7 and caspase-8 measured by luminescent assays; (ii) at low concentrations, PUAs and HEPEs affected the expression levels of caspase-8 and caspase-3/7 (isolated for the first time here in P. lividus) genes, detected by Real Time qPCR. These findings have interesting implications from the ecological point of view, given the importance of diatom blooms in nutrient-rich aquatic environments.

  12. Activation of caspase 8 in the pituitaries of streptozotocin-induced diabetic rats: implication in increased apoptosis of lactotrophs.

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    Arroba, Ana I; Frago, Laura M; Argente, Jesús; Chowen, Julie A

    2005-10-01

    Lactotroph cell death is increased in streptozotocin-induced diabetic rats. To determine the mechanism involved, cell death proteins were accessed in pituitaries of diabetic (streptozotocin at 65 mg/kg, 2 months evolution) and control male rats by Western blot analysis and double immunohistochemistry. The intact and cleaved forms of caspase 9 were increased in diabetic rat pituitaries compared with controls. Although the proforms of caspases 3, 6, and 7 were increased in diabetic rat pituitaries, their activated forms were either unchanged or decreased. Activation of these effector caspases may be blocked by the increased expression of X-chromosome-linked inhibitor of apoptosis protein (XIAP) in diabetic rat pituitaries. However, in diabetic rats, XIAP expression in lactotrophs was decreased, suggesting that this cell type is not protected. Caspase 8, p53, and nuclear factor kappaB were more highly activated in diabetic rat pituitaries, with caspase 8 colocalization in lactotrophs being increased. These results suggest that, in the pituitaries of diabetic rats, the cascades of normal cell turnover are partially inhibited, possibly via XIAP, and this may be cell specific. Furthermore, activation of the extrinsic cell-death pathway, including activation of caspase 8, may underlie the diabetes-associated increase in lactotroph death.

  13. Staphylococcus aureus - induced tumor necrosis factor - related apoptosis - inducing ligand expression mediates apoptosis and caspase-8 activation in infected osteoblasts

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    Bost Kenneth L

    2003-04-01

    Full Text Available Abstract Background Staphylococcus aureus infection of normal osteoblasts induces expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL. Results Normal osteoblasts were incubated in the presence of purified bacterial products over a range of concentrations. Results demonstrate that purified surface structures and a selected superantigen present in the extracellular environment are not capable of inducing TRAIL expression by osteoblasts. Osteoblasts were co-cultured with S. aureus at various multiplicities of infection utilizing cell culture chamber inserts. Results of those experiments suggest that direct contact between bacteria and osteoblasts is necessary for optimal TRAIL induction. Finally, S. aureus infection of osteoblasts in the presence of anti-TRAIL antibody demonstrates that TRAIL mediates caspase-8 activation and apoptosis of infected cells. Conclusions Collectively, these findings suggest a mechanism whereby S. aureus mediates bone destruction via induction of osteoblast apoptosis.

  14. Irciniastatin A induces JNK activation that is involved in caspase-8-dependent apoptosis via the mitochondrial pathway.

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    Chinen, Takumi; Nagumo, Yoko; Watanabe, Tsubasa; Imaizumi, Takamichi; Shibuya, Masatoshi; Kataoka, Takao; Kanoh, Naoki; Iwabuchi, Yoshiharu; Usui, Takeo

    2010-12-15

    Irciniastatin A (ISA)/psymberin, a pederin-type natural product isolated from marine sponge, exhibits extremely potent and selective cytotoxicity against certain human cancer cell lines, but its molecular target and cytotoxic mechanisms are still unknown. Here we show that ISA is a potent inhibitor of protein translation, and induces apoptosis accompanied with activation of the stress-activated protein kinases via the mitochondrial pathway in human leukemia Jurkat cells. ISA potently inhibited protein translation, and induced a slow but prolonged activation of the stress-activated protein kinases, JNK and p38, at between 1h and 6h after treatment. In Bcl-x(L)-transfected cells, the activation of JNK and p38 by ISA was shortened. The same results were obtained in the cells treated with N-acetyl-L-cysteine, suggesting that the prolonged activation of JNK and p38 by ISA is mediated by reactive oxygen species generated from mitochondria. ISA strongly induced apoptosis, which was partially suppressed by the JNK inhibitor SP600125, but not by the p38 inhibitor SB202190. Apoptosis induction by ISA was partially reduced, but not suppressed by SP600125 in caspase-8-deficient Jurkat cells. These results suggest that ISA activates stress-activated kinases by a mitochondria-mediated mechanism, and that activation of JNK is required for caspase-8-dependent apoptosis.

  15. Fas-Induced Apoptosis of Renal Cell Carcinoma is Mediated by Apoptosis Signal-Regulating Kinase 1 via Mitochondrial Damage-Dependent Caspase-8 Activation

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    Mohamed Hassan

    2009-01-01

    Full Text Available Renal cell carcinoma (RCC is a prototype of a chemo refractory tumour. It remains the most lethal of the common urologic cancers and is highly resistant to conventional therapy. Here, we confirmed the efficiency of anti-Fas monoclonal antibody (CH11 as alternative therapeutic approach for the treatment of RCC and investigated the molecular mechanism(s, whereby CH11 induces apoptosis of RCC cells. The present study shows an essential role for apoptosis signal-regulating kinase 1 (ASK1, together with both c-jun-N-terminal kinase (JNK and p38 pathways, and caspase-8 in this process. Furthermore, CH11-dependent induction of the ASK1–JNK/p38 pathways was found to activate the transcription factors AP-1 and ATF-2, and FADD-caspase-8-Bid signalling, resulting in the translocation of both Bax and Bak proteins, and subsequently mitochondrial dysregulation that is characterized by the loss of mitochondrial membrane potential (ΔΨm, cytochrome c release and cleavage of caspase-9, caspase-3 and PARP. Thus, the described molecular mechanisms of CH11-induced apoptosis suggest the reliability of Fas activation as an alternative therapeutic approach for the treatment of patients with advanced renal cell carcinoma.

  16. Combined treatment with the Cox-2 inhibitor niflumic acid and PPARγ ligand ciglitazone induces ER stress/caspase-8-mediated apoptosis in human lung cancer cells.

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    Kim, Byeong Mo; Maeng, Kyungah; Lee, Kee-Ho; Hong, Sung Hee

    2011-01-28

    The present study was performed to investigate the possible combined use of the Cox-2 inhibitor niflumic acid and the PPARγ ligand ciglitazone and to elucidate the mechanisms underlying enhanced apoptosis by this combination treatment in human lung cancer cells. Combined niflumic acid-ciglitazone treatment synergistically induced apoptotic cell death, activated caspase-9, caspase-3, and induced caspase-3-mediated PARP cleavage. The combination treatment also triggered apoptosis through caspase-8/Bid/Bax activation, and the inhibition of caspase-8 suppressed caspase-8/Bid activation, caspase-3-mediated PARP cleavage, and concomitant apoptosis. In addition, combined niflumic acid-ciglitazone treatment significantly induced ER stress responses, and suppression of CHOP expression significantly attenuated the combined niflumic acid-ciglitazone treatment-induced activation of caspase-8 and caspase-3, and the subsequent apoptotic cell death, indicating a role of ER stress in caspase-8 activation and apoptosis. Interestingly, the pro-apoptotic effects of combined niflumic acid-ciglitazone treatment were realized through Cox-2- and PPARγ-independent mechanisms. Taken together, these results suggest that sequential ER stress and caspase-8 activation are critical in combined niflumic acid-ciglitazone treatment-induced apoptosis in human lung cancer cells.

  17. Neuronal deletion of caspase 8 protects against brain injury in mouse models of controlled cortical impact and kainic acid-induced excitotoxicity.

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    Maryla Krajewska

    Full Text Available Acute brain injury is an important health problem. Given the critical position of caspase 8 at the crossroads of cell death pathways, we generated a new viable mouse line (Ncasp8(-/-, in which the gene encoding caspase 8 was selectively deleted in neurons by cre-lox system.Caspase 8 deletion reduced rates of neuronal cell death in primary neuronal cultures and in whole brain organotypic coronal slice cultures prepared from 4 and 8 month old mice and cultivated up to 14 days in vitro. Treatments of cultures with recombinant murine TNFα (100 ng/ml or TRAIL (250 ng/mL plus cyclohexamide significantly protected neurons against cell death induced by these apoptosis-inducing ligands. A protective role of caspase 8 deletion in vivo was also demonstrated using a controlled cortical impact (CCI model of traumatic brain injury (TBI and seizure-induced brain injury caused by kainic acid (KA. Morphometric analyses were performed using digital imaging in conjunction with image analysis algorithms. By employing virtual images of hundreds of brain sections, we were able to perform quantitative morphometry of histological and immunohistochemical staining data in an unbiased manner. In the TBI model, homozygous deletion of caspase 8 resulted in reduced lesion volumes, improved post-injury motor performance, superior learning and memory retention, decreased apoptosis, diminished proteolytic processing of caspases and caspase substrates, and less neuronal degeneration, compared to wild type, homozygous cre, and caspase 8-floxed control mice. In the KA model, Ncasp8(-/- mice demonstrated superior survival, reduced seizure severity, less apoptosis, and reduced caspase 3 processing. Uninjured aged knockout mice showed improved learning and memory, implicating a possible role for caspase 8 in cognitive decline with aging.Neuron-specific deletion of caspase 8 reduces brain damage and improves post-traumatic functional outcomes, suggesting an important role for this

  18. Essential role of caspase-8 in p53/p73-dependent apoptosis induced by etoposide in head and neck carcinoma cells

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    Uematsu Hiroshi

    2011-07-01

    Full Text Available Abstract Background Caspase-8 is a key upstream mediator in death receptor-mediated apoptosis and also participates in mitochondria-mediated apoptosis via cleavage of proapoptotic Bid. However, the role of caspase-8 in p53- and p73-dependent apoptosis induced by genotoxic drugs remains unclear. We recently reported that the reconstitution of procaspase-8 is sufficient for sensitizing cisplatin- but not etoposide-induced apoptosis, in chemoresistant and caspase-8 deficient HOC313 head and neck squamous cell carcinoma (HNSCC cells. Results We show that p53/p73-dependent caspase-8 activation is required for sensitizing etoposide-induced apoptosis by utilizing HOC313 cells carrying a temperature-sensitive p53G285K mutant. Restoration of wild-type p53 function under the permissive conditions, together with etoposide treatment, led to substantial transcriptional activation of proapoptotic Noxa and PUMA, but failed to induce apoptosis. In addition to p53 restoration, caspase-8 reconstitution was needed for sensitization to etoposide-induced apoptosis, mitochondria depolarization, and cleavage of the procaspases-3, and -9. In etoposide-sensitive Ca9-22 cells carrying a temperature-insensitive mutant p53, siRNA-based p73 knockdown blocked etoposide-induced apoptosis and procaspase-8 cleavage. However, induction of p73 protein and up-regulation of Noxa and PUMA, although observed in Ca9-22 cells, were hardly detected in etoposide-treated HOC313 cells under non-permissive conditions, suggesting a contribution of p73 reduction to etoposide resistance in HOC313 cells. Finally, the caspase-9 inhibitor Ac-LEHD-CHO or caspase-9 siRNA blocked etoposide-induced caspase-8 activation, Bid cleavage, and apoptosis in both cell lines, indicating that p53/p73-dependent caspase-8 activation lies downstream of mitochondria. Conclusions we conclude that p53 and p73 can act as upstream regulators of caspase-8, and that caspase-8 is an essential mediator of the p53/p73

  19. Compound K, a metabolite of ginseng saponin, induces apoptosis via caspase-8-dependent pathway in HL-60 human leukemia cells

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    Choi Jung-Hye

    2009-12-01

    Full Text Available Abstract Background Compound K [20-O-β-(D-glucopyranosyl-20(S-protopanaxadiol], a metabolite of the protopanaxadiol-type saponins of Panax ginseng C.A. Meyer, has been reported to possess anti-tumor properties to inhibit angiogenesis and to induce tumor apoptosis. In the present study, we investigated the effect of Compound K on apoptosis and explored the underlying mechanisms involved in HL-60 human leukemia cells. Methods We examined the effect of Compound K on the viabilities of various cancer cell lines using MTT assays. DAPI assay, Annexin V and PI double staining, Western blot assay and immunoprecipitation were used to determine the effect of Compound K on the induction of apoptosis. Results Compound K was found to inhibit the viability of HL-60 cells in a dose- and time-dependent manner with an IC50 of 14 μM. Moreover, this cell death had typical features of apoptosis, that is, DNA fragmentation, DNA ladder formation, and the externalization of Annexin V targeted phosphatidylserine residues in HL-60 cells. In addition, compound-K induced a series of intracellular events associated with both the mitochondrial- and death receptor-dependent apoptotic pathways, namely, (1 the activation of caspases-3, -8, and -9; (2 the loss of mitochondrial membrane potential; (3 the release of cytochrome c and Smac/DIABLO to the cytosol; (4 the translocation of Bid and Bax to mitochondria; and (5 the downregulations of Bcl-2 and Bcl-xL. Furthermore, a caspase-8 inhibitor completely abolished caspase-3 activation, Bid cleavage, and subsequent DNA fragmentation by Compound K. Interestingly, the activation of caspase-3 and -8 and DNA fragmentation were significantly prevented in the presence of cycloheximide, suggesting that Compound K-induced apoptosis is dependent on de novo protein synthesis. Conclusions The results indicate that caspase-8 plays a key role in Compound K-stimulated apoptosis via the activation of caspase-3 directly or indirectly through

  20. Caspase-8 Activation Precedes Alterations of Mitochondrial Membrane Potential during Monocyte Apoptosis Induced by Phagocytosis and Killing of Staphylococcus aureus

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    Węglarczyk, Kazimierz; Baran, Jarosław; Zembala, Marek; Pryjma, Juliusz

    2004-01-01

    Human peripheral blood monocytes become apoptotic following phagocytosis and killing of Staphylococcus aureus. Although this type of monocyte apoptosis is known to be initiated by Fas-Fas ligand (FasL) interactions, the downstream signaling pathway has not been determined. In this work the involvement of mitochondria and the kinetics of caspase-8 and caspase-3 activation after phagocytosis of S. aureus were studied. Caspase-8 activity was measured in cell lysates by using the fluorogenic substrate Ac-IETD-AFC. Active caspase-3 levels and mitochondrial membrane potential (Δψm) were measured in whole cells by flow cytometry using monoclonal antibodies reacting with activated caspase-3 and chloromethyl-X-rosamine, respectively. The results show that caspase-8 was activated shortly after phagocytosis of bacteria. Caspase-8 activation was followed by progressive disruption of Δψm, which is associated with the production of reactive oxygen intermediates. The irreversible caspase-8 inhibitor zIETD-FMK prevented the disruption of Δψm and the release of cytochrome c from S. aureus-exposed monocytes. Caspase-3 activation occurred following disruption of Δψm. These results strongly suggest that apoptosis of monocytes that have phagocytosed and killed S. aureus is driven by the Fas-FasL-initiated pathway, which is typical for type II cells. PMID:15102767

  1. Smac Mimetic Bypasses Apoptosis Resistance in FADD- or Caspase-8-Deficient Cells by Priming for Tumor Necrosis Factor α-Induced Necroptosis

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    Bram Laukens

    2011-10-01

    Full Text Available Searching for new strategies to bypass apoptosis resistance, we investigated the potential of the Smac mimetic BV6 in Jurkat leukemia cells deficient in key molecules of the death receptor pathway. Here, we demonstrate for the first time that Smac mimetic primes apoptosis-resistant, FADD- or caspase-8-deficient leukemia cells for TNFα-induced necroptosis in a synergistic manner. In contrast to TNFα, Smac mimetic significantly enhances CD95-induced apoptosis in wild-type but not in FADD-deficient cells. Interestingly, Smac mimetic- and TNFα-mediated cell death occurs without characteristic features of apoptosis (i.e., caspase activation, DNA fragmentation in FADD-deficient cells. By comparison, Smac mimetic and TNFα trigger activation of caspase-8, -9, and -3 and DNA fragmentation in wild-type cells. Consistently, the caspase inhibitor zVAD.fmk fails to block Smac mimetic- and TNFα-triggered cell death in FADD- or caspase-8-deficient cells, while it confers protection in wild-type cells. By comparison, necrostatin-1, an RIP1 kinase inhibitor, abolishes Smac mimetic- and TNFα-induced cell death in FADD- or caspase-8-deficient. Thus, Smac mimetic enhances TNFα-induced cell death in leukemia cells via two distinct pathways in a context-dependent manner: it primes apoptosis-resistant cells lacking FADD or caspase-8 to TNFα-induced, RIP1-dependent and caspase-independent necroptosis, whereas it sensitizes apoptosis-proficient cells to TNFα-mediated, caspase-dependent apoptosis. These findings have important implications for the therapeutic exploitation of necroptosis as an alternative cell death program to overcome apoptosis resistance.

  2. Upregulation and activation of caspase-3 or caspase-8 and elevation of intracellular free calcium mediated apoptosis of indomethacin-induced K562 cells

    Institute of Scientific and Technical Information of China (English)

    张广森; 周光飚; 戴崇文

    2004-01-01

    Background A nonsteroidal anti-inflammatory drug, indomethacin, has been shown to have anti-leukemic activity and induce leukemic cell opoptosis. This study was to elucidate the mechanism of indomethacin-induced K562 cell apoptosis.Methods K562 cells were grown in RPMI 1640 medium and treated with different doses of indomethacin (0 μmol/L, 100 μmol/L, 200 μmol/L, 400 μmol/L, 800 μmol/L) for 72 hours. The cells were harvested, and cell viability or apoptosis was analyzed using MTT assay and AO/EB stain, combining laser scanning confocal microscopy (LSCM) technique separately. For the localization and distribution of intracellular caspase-3 or caspase-8 protein, immunofluorescence assay was carried out. To reveal the activation of caspase-3 or caspase-8 in indomethacin-treated cells, Western blot detection was used. The change in intracellular free calcium was determined by Fluo-3/ Am probe labeling combined with LSCM. Results Indomethacin could lead to K562 cell apoptosis and inhibit cell viability in a concentration-dependent manner. An increased expression of intracellular caspase-3 or caspase-8 was observed at higher doses of indomethacin (400-800 μmol/L). Western blot results showed upregrulation and activation in both caspase-3 and caspase-8 protein. Under indomethacin intervention, the levels of intracellular free calcium showed a significant increase. Blocking the activity of cyclooxygenase did not abolish the effects of indomethacin on K562 cell apoptosis.Conclusions Activation and upregulation of caspase-3 or caspase-8 protein were responsible for Indomethacin-induced K562 cell apoptosis. Variation of intracellular free calcium might switch on the apoptotic pathway and the proapoptotic effect of indomethacin might be cyclooxygenase-independent.

  3. Standardized bioactive fraction of Phaleria macrocarpa(Proliverenol) prevents ethanol-induced hepatotoxicity via down-regulation of NF-kB-TNFα-caspase-8 pathway

    Institute of Scientific and Technical Information of China (English)

    Guntur Berlian; Olivia Mayasari Tandrasasmita; Raymond Rubianto Tjandrawinata

    2016-01-01

    Objective: To verify that Proliverenol has a potential ability in protecting cells from ethanol-induced hepatotoxicity.Methods: Activity of Proliverenol against ethanol-induced apoptosis was evaluated at m RNA and protein levels in Hep G2 cell exposed to Proliverenol for 1 and 3 h.Results: Proliverenol conferred hepatoprotective activity through increasing cell survival up to 53%–69% via up-regulation of APEX1 DNA repair enzyme for 3.0–4.7 fold and down-regulating of nuclear factor-kB, tumor necrosis factora and caspase-8 expression,allowing them to prevent 4.5–6.9 fold of alanine aminotransferase(ALT) leakage in Hep G2 cells. Our finding revealed that Proliverenol repressed expression of ALT, which is significantly important as possible alternative mechanism for increased blood transaminase activities. In addition, the result also showed that caspase-8 pathway seemed to be involved in the molecular pathway rather than directly inducing mitochondrial damage.Conclusions: The data support our hypothesis that Proliverenol has a potential ability in protecting cells from ethanol-induced hepatotoxicity. We propose that Proliverenol provides hepatoprotective activity through up-regulating expression of APEX1 that repress DNA fragmentation, and down-regulating expression of nuclear factor-kB, tumor necrosis factora and caspase-8, which therefore repress ALT leakage and its expression.

  4. Pro-apoptotic Chemotherapeutic Drugs Induce Non-canonical Processing and Release of IL-1β via Caspase-8 in Dendritic Cells#

    OpenAIRE

    Antonopoulos, Christina; El Sanadi, Caroline; Kaiser, William J.; Mocarski, Edward S.; Dubyak, George R.

    2013-01-01

    The identification of non-canonical (caspase-1 independent) pathways for IL-1β production has unveiled an intricate interplay between inflammatory and death-inducing signaling platforms. We found a heretofore unappreciated role for caspase-8 as a major pathway for IL-1β processing and release in murine bone marrow-derived dendritic cells (BMDC) co-stimulated with TLR4 agonists and pro-apoptotic chemotherapeutic agents such as doxorubicin (Dox) or staurosporine (STS). The ability of Dox to sti...

  5. pERK 1/2 inhibit Caspase-8 induced apoptosis in cancer cells by phosphorylating it in a cell cycle specific manner.

    Science.gov (United States)

    Mandal, Ranadip; Raab, Monika; Matthess, Yves; Becker, Sven; Knecht, Rainald; Strebhardt, Klaus

    2014-03-01

    ERK 1/2 are found to be hyperactive in many cancers. Active ERK 1/2 (pERK 1/2) are known to protect cancer cells from undergoing death receptor-mediated apoptosis, although the mechanism(s) behind this is poorly understood. Through in vitro kinase assays and mass-spectrometry we demonstrate that pERK 1/2 can phosphorylate pro-Caspase-8 at S387. Also, in EGFR-overexpressing Type I and II ovarian and breast cancer cell lines respectively, ERK 1/2 remain active only during the interphase. During this period, pERK 1/2 could inhibit Trail-induced apoptosis, most effectively during the G1/S phase. By knocking-down the endogenous pro-Caspase-8 using RNAi and replacing it with its non-phosphorylatable counterpart (S387A), a significant increase in Caspase-8 activity upon Trail stimulation was observed, even in the presence of pERK 1/2. Taken together, we propose that a combination of Trail and an inhibitor of ERK 1/2 activities could potentially enhance of Trail's effectiveness as an anti-cancer agent in ERK 1/2 hyperactive cancer cells.

  6. Proteasomal regulation of caspase-8 in cancer cell apoptosis.

    Science.gov (United States)

    Fiandalo, Michael V; Schwarze, Steven R; Kyprianou, Natasha

    2013-06-01

    Previous studies demonstrated that proteasome inhibition sensitizes TRAIL resistant prostate cancer cells to TRAIL-mediated apoptosis via stabilization of the active p18 subunit of caspase-8. The present study investigated the impact of proteasome inhibition on caspase-8 stability, ubiquitination, trafficking, and activation in cancer cells. Using caspase-8 deficient neuroblastoma (NB7) cells for reconstituting non-cleavable mutant forms of caspase-8, we demonstrated that the non-cleavable forms of caspase-8 are capable of inducing apoptosis comparably to wild-type caspase-8, in response to proteasome inhibitor and GST-TRAIL. Moreover in the LNCaP human prostate cancer cells, caspase-8 polyubiquitination occurs after TRAIL stimulation and caspase-8 processing. Subcellular fractionation analysis revealed caspase-8 activity in both cytosol and plasma membrane fractions in both NB7 reconstituted caspase-8 cell lines, as well the LNCaP prostate cancer cells. The present results suggest that caspase-8 stabilization through proteasome inhibition leads to reactivation of the extrinsic pathway of apoptosis and identify E3 ligase mediating caspase-8 polyubiquitination, as a novel molecular target. Inhibition of this E3 ligase in combination with TRAIL towards restoring apoptosis signaling activation may have potential therapeutic significance in resistant tumors.

  7. Association of active caspase 8 with the mitochondrial membrane during apoptosis: potential roles in cleaving BAP31 and caspase 3 and mediating mitochondrion-endoplasmic reticulum cross talk in etoposide-induced cell death.

    Science.gov (United States)

    Chandra, Dhyan; Choy, Grace; Deng, Xiaodi; Bhatia, Bobby; Daniel, Peter; Tang, Dean G

    2004-08-01

    It was recently demonstrated that during apoptosis, active caspase 9 and caspase 3 rapidly accumulate in the mitochondrion-enriched membrane fraction (D. Chandra and D. G. Tang, J. Biol. Chem.278:17408-17420, 2003). We now show that active caspase 8 also becomes associated with the membranes in apoptosis caused by multiple stimuli. In MDA-MB231 breast cancer cells treated with etoposide (VP16), active caspase 8 is detected only in the membrane fraction, which contains both mitochondria and endoplasmic reticulum (ER), as revealed by fractionation studies. Immunofluorescence microscopy, however, shows that procaspase 8 and active caspase 8 predominantly colocalize with the mitochondria. Biochemical analysis demonstrates that both procaspase 8 and active caspase 8 are localized mainly on the outer mitochondrial membrane (OMM) as integral proteins. Functional analyses with dominant-negative mutants, small interfering RNAs, peptide inhibitors, and Fas-associated death domain (FADD)- and caspase 8-deficient Jurkat T cells establish that the mitochondrion-localized active caspase 8 results mainly from the FADD-dependent and tumor necrosis factor receptor-associated death domain-dependent mechanisms and that caspase 8 activation plays a causal role in VP16-induced caspase 3 activation and cell death. Finally, we present evidence that the OMM-localized active caspase 8 can activate cytosolic caspase 3 and ER-localized BAP31. Cleavage of BAP31 leads to the generation of ER- localized, proapoptotic BAP20, which may mediate mitochondrion-ER cross talk through a Ca(2+)-dependent mechanism.

  8. Drug-induced caspase 8 upregulation sensitises cisplatin-resistant ovarian carcinoma cells to rhTRAIL-induced apoptosis

    NARCIS (Netherlands)

    Duiker, E. W.; Meijer, A.; van der Bilt, A. R. M.; Meersma, G. J.; Kooi, N.; van der Zee, A. G. J.; de Vries, E. G.; de Jong, S.

    2011-01-01

    BACKGROUND: Drug resistance is a major problem in ovarian cancer. Triggering apoptosis using death ligands such as tumour necrosis factor-related apoptosis inducing ligand (TRAIL) might overcome chemoresistance. METHODS: We investigated whether acquired cisplatin resistance affects sensitivity to re

  9. Molecular cloning, immunohistochemical localization, characterization and expression analysis of caspase-8 from the blunt snout bream (Megalobrama amblycephala) exposed to ammonia.

    Science.gov (United States)

    Sun, Shengming; Ge, Xianping; Zhu, Jian; Zhang, Wuxiao; Zhang, Qiong

    2015-12-01

    Caspase-8 is an initiator caspase that plays a crucial role in some cases of apoptosis by extrinsic and intrinsic pathways. Caspase-8 structure and function have been extensively studied in mammals, but in fish the characterization of that initiator caspase is still scarce. In this study, we isolated the caspase-8 gene from Megalobrama amblycephala, one of the most important industrial aquatic animals in China using rapid amplification of cDNA ends (RACE). The 2034 bp full-length M. amblycephala caspase-8 cDNA sequence contained an ORF of 1467 bp encoding a polypeptide of 489 amino acid residues, a 5'-UTR of 102 bp and a 3'-UTR of 462 bp. The caspase-8 amino acid sequences contained two highly conservative death effector domains (DEDs) at N-terminal, the caspase family domains P20 and P10, caspase-8 active-site pentapeptide and potential aspartic acid cleavage sites. Phylogenetic analysis revealed that M. amblycephala caspase-8 were clustered with the caspase-8 from other vertebrate. Real-time quantitative PCR analysis revealed that caspase-8 transcripts were detected in liver after exposure to ammonia. Meanwhile using Western blot analysis, caspase-8 cleaved fragment was detected and significant alteration of procaspase-8 level was found with the same ammonia treatment condition. Furthermore, the result of immunohistochemical detection showed that remarkable changes of immunopositive staining were observed after ammonia treatment. Accordingly, the results signify that caspase-8 of fish may play an essential role in ammonia induced apoptosis.

  10. Docosahexaenoic acid induces apoptosis in MCF-7 cells in vitro and in vivo via reactive oxygen species formation and caspase 8 activation.

    Directory of Open Access Journals (Sweden)

    Ki Sung Kang

    Full Text Available BACKGROUND: The present study sought to further investigate the in vitro and in vivo anticancer effects of a representative omega-3 fatty acid, docosahexaenoic acid (DHA, with a focus on assessing the induction of oxidative stress and apoptosis as an important mechanism for its anticancer actions. METHODOLOGY/PRINCIPAL FINDINGS: In vitro studies showed that DHA strongly reduces the viability and DNA synthesis of MCF-7 human breast cancer cells in culture, and also promotes cell death via apoptosis. Mechanistically, accumulation of reactive oxygen species and activation of caspase 8 contribute critically to the induction of apoptotic cell death. Co-presence of antioxidants or selective inhibition or knockdown of caspase 8 each effectively abrogates the cytotoxic effect of DHA. Using athymic nude mice as an in vivo model, we found that feeding animals the 5% fish oil-supplemented diet for 6 weeks significantly reduces the growth of MCF-7 human breast cancer cells in vivo through inhibition of cancer cell proliferation as well as promotion of cell death. Using 3-nitrotyrosine as a parameter, we confirmed that the fish oil-supplemented diet significantly increases oxidative stress in tumor cells in vivo. Analysis of fatty acid content in plasma and tissues showed that feeding animals a 5% fish oil diet increases the levels of DHA and eicosapentaenoic acid in both normal and tumorous mammary tissues by 329% and 300%, respectively. CONCLUSIONS/SIGNIFICANCE: DHA can strongly induce apoptosis in human MCF-7 breast cancer cells both in vitro and in vivo. The induction of apoptosis in these cells is selectively mediated via caspase 8 activation. These observations call for further studies to assess the effectiveness of fish oil as a dietary supplement in the prevention and treatment of human breast cancer.

  11. Andrographolide enhances 5-fluorouracil-induced apoptosis via caspase-8-dependent mitochondrial pathway involving p53 participation in hepatocellular carcinoma (SMMC-7721) cells.

    Science.gov (United States)

    Yang, Lu; Wu, Dingfang; Luo, Kewang; Wu, Shihua; Wu, Ping

    2009-04-18

    Despite recent significant advances in the treatment of human carcinoma (HCC), the results of chemotherapy to date remain unsatisfactory. 5-Fluorouracil (5-FU) still represents the cornerstone of treatment of carcinoma, and resistance to the actions of 5-FU is a major obstacle to successful chemotherapy. More effective treatment strategies may involve combinations of agents with activity against HCC. Andrographolide (ANDRO), a natural bicyclic diterpenoid lactone isolated from Andrographis paniculata, has been shown to suppress the growth of HCC cells and trigger apoptosis in vitro. To assess the suitability of ANDRO as a chemotherapeutic agent in HCC, its cytotoxic effects have been evaluated both as a single agent and in combination with 5-FU. ANDRO potentiates the cytotoxic effect of 5-FU in HCC cell line SMMC-7721 through apoptosis. ANDRO alone induces SMMC-7721 apoptosis with p53 expression, Bax conformation and caspase-3,8,9 activation. Surprisingly, the addition of ANDRO to 5-FU induces synergistic apoptosis, which could be corroborated to the increased caspase-8, p53 activity and the significant changes of Bax conformation in these cells, resulting in increased losses of mitochondrial membrane potential, increased release of cytochrome c, and activation of caspase-9 and caspase-3. Suppression of caspase-8 with the specific inhibitor z-IETD-fmk abrogates largely ANDRO/5-FU biological activity by preventing mitochondrial membrane potential disappearance, caspase-3,9 activation and subsequent apoptosis. The results suggest that ANDRO may be effective in combination with 5-FU for the treatment of HCC cells SMMC-7721.

  12. Artemisinin induces caspase-8/9-mediated and Bax/Bak-independent apoptosis in human lung adenocarcinoma (ASTC-a-1) cells.

    Science.gov (United States)

    Xiao, Feng-Lian; Gao, Wei-Jie; Liu, Cheng-Yi; Wang, Xiao-Ping; Chen, Tong-Sheng

    2011-01-01

    Artemisinin (ARTE), an antimalarial phytochemical component from the sweet wormwood plant, has been shown a potential anticancer activity by inducing cell apoptosis. The aim of this report is to explore the mechanism of ARTE-induced human lung adenocarcinoma (ASTC-a-1) cell apoptosis. Cell counting kit (CCK-8) assay showed that ARTE induced cytotoxcity in a dose- and time-dependent manner. Confocal microscopy fluorescence imaging of cells stained with Hoechst 33258 and flow cytometry (FCM) analysis of cells stained with Annexin V-FITC/propidium iodide (PI) showed that ARTE induced reactive oxygen species (ROS)-dependent apoptosis. Confocal fluorescence resonance energy transfer (FRET) imaging of single living cells expressing SCAT3, SCAT9 or CFP-Bid-YFP and fluorometic substrate assay showed that ARTE induced the activation of caspase-3, -8 and -9. Moreover, inhibition of caspase-8 or -9 completely blocked ARTE-induced apoptosis which was only partially attenuated by caspase-3 inhibitor. Interestingly, silencing Bax and Bak by RNA interference (RNAi) did not attenuate ARTE-induced apoptosis. Collectively, ARTE induces caspase-dependent but Bax/Bak-independent apoptosis in ASTC-a-1 cells.

  13. Metallic nickel nano- and fine particles induce JB6 cell apoptosis through a caspase-8/AIF mediated cytochrome c-independent pathway

    Directory of Open Access Journals (Sweden)

    Castranova Vincent

    2009-04-01

    Full Text Available Abstract Background Carcinogenicity of nickel compounds has been well documented. However, the carcinogenic effect of metallic nickel is still unclear. The present study investigates metallic nickel nano- and fine particle-induced apoptosis and the signal pathways involved in this process in JB6 cells. The data obtained from this study will be of benefit for elucidating the pathological and carcinogenic potential of metallic nickel particles. Results Using 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay, we found that metallic nickel nanoparticles exhibited higher cytotoxicity than fine particles. Both metallic nickel nano- and fine particles induced JB6 cell apoptosis. Metallic nickel nanoparticles produced higher apoptotic induction than fine particles. Western-blot analysis showed an activation of proapoptotic factors including Fas (CD95, Fas-associated protein with death domain (FADD, caspase-8, death receptor 3 (DR3 and BID in apoptotic cells induced by metallic nickel particles. Immunoprecipitation (IP western blot analysis demonstrated the formation of the Fas-related death-inducing signaling complex (DISC in the apoptotic process. Furthermore, lamin A and beta-actin were cleaved. Moreover, we found that apoptosis-inducing factor (AIF was up-regulated and released from mitochondria to cytoplasm. Interestingly, although an up-regulation of cytochrome c was detected in the mitochondria of metallic nickel particle-treated cells, no cytochrome c release from mitochondria to cytoplasm was found. In addition, activation of antiapoptotic factors including phospho-Akt (protein kinase B and Bcl-2 was detected. Further studies demonstrated that metallic nickel particles caused no significant changes in the mitochondrial membrane permeability after 24 h treatment. Conclusion In this study, metallic nickel nanoparticles caused higher cytotoxicity and apoptotic induction than fine particles in JB6 cells. Apoptotic cell death

  14. The Role of the Caspase-8 Inhibitor FLIP in Androgen-Withdrawal Induced Death of Prostate Epithelium

    Science.gov (United States)

    2007-01-01

    precipitation and immunoblotting to measure FLIP protein levels in apoptosing rat prostate glands. • 1.2 Use immunohistochemistry and in situ hybridization to...determine the spatial (cell-type specific) pattern of FLIP expression in apoptosing rat prostate glands. • 1.3 Measure the levels of FLIP mRNA (by...quantitative RT-PCR) and protein (by affinity precipitation and immunoblotting) in NRP-152 cells induced to apoptose by TGFß. Results: We previously

  15. Id3 induces an Elk-1–caspase-8-dependent apoptotic pathway in squamous carcinoma cells

    OpenAIRE

    Chen, You-Shin; Aubee, Joseph; Divito, Kyle A.; Zhou, Hengbo; Zhang, Weiyi; Chou, Fen-Pi; Simbulan-Rosenthal, Cynthia M.; Rosenthal, Dean S.

    2015-01-01

    Inhibitor of differentiation/DNA-binding (Id) proteins are helix–loop–helix (HLH) transcription factors. The Id protein family (Id1–Id4) mediates tissue homeostasis by regulating cellular processes including differentiation, proliferation, and apoptosis. Ids typically function as dominant negative HLH proteins, which bind other HLH proteins and sequester them away from DNA promoter regions. Previously, we have found that Id3 induced apoptosis in immortalized human keratinocytes upon UVB expos...

  16. Autoregulatory Feedback Mechanism of P38MAPK/Caspase-8 in Photodynamic Therapy-Hydrophilic/Lipophilic Tetra-α-(4-carboxyphenoxy Phthalocyanine Zinc-Induced Apoptosis of Human Hepatocellular Carcinoma Bel-7402 Cells

    Directory of Open Access Journals (Sweden)

    Yu Wang

    2014-01-01

    Full Text Available Photodynamic therapy (PDT is a novel and promising antitumor treatment. Our previous study showed that hydrophilic/lipophilic tetra-α-(4-carboxyphenoxy phthalocyanine zinc- (TαPcZn- mediated PDT (TαPcZn-PDT inhibits the proliferation of human hepatocellular carcinoma Bel-7402 cells by triggering apoptosis and arresting cell cycle. However, mechanisms of TαPcZn-PDT-induced apoptosis of Bel-7402 cells have not been fully clarified. In the present study, therefore, effect of TαPcZn-PDT on apoptosis, P38MAPK, p-P38MAPK, Caspase-8, Caspase-3, Bcl-2, Bid, Cytochrome c, and mitochondria membrane potential in Bel-7402 cells without or with P38MAPK inhibitor SB203580 or Caspase-8 inhibitor Ac-IEFD-CHO was investigated by haematoxylin and eosin (HE staining assay, flow cytometry analysis of annexin V-FITC/propidium iodide (PI double staining cells and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide (JC-1, and immunoblot assay. We found that TαPcZn-PDT resulted in apoptosis induction, activation of P38MAPK, Caspase-8, Caspase-3, and Bid, downregulation of Bcl-2, release of Cytochrome c from mitochondria, and disruption of mitochondrial membrane potential in TαPcZn-PDT-treated Bel-7402 cells. In contrast, SB203580 or Ac-IEFD-CHO attenuated induction of apoptosis, activation of P38MAPK, Caspase-8, Caspase-3, and Bid, downregulation of Bcl-2, release of Cytochrome c from mitochondria, and disruption of mitochondrial membrane potential in TαPcZn-PDT-treated Bel-7402 cells. Taken together, we conclude that Caspase-3, Bcl-2, Bid, and mitochondria are involved in autoregulatory feedback of P38MAPK/Caspase-8 during TαPcZn-PDT-induced apoptosis of Bel-7402 cells.

  17. Alterations in blood pressure, antioxidant status and caspase 8 expression in cobalt chloride-induced cardio-renal dysfunction are reversed by Ocimum gratissimum and gallic acid in Wistar rats.

    Science.gov (United States)

    Akinrinde, A S; Oyagbemi, A A; Omobowale, T O; Asenuga, E R; Ajibade, T O

    2016-07-01

    The protective abilities of the chloroform extract of Ocimum gratissimum (COG) and gallic acid against cobalt chloride (CoCl2) - induced cardiac and renal toxicity were evaluated. Rats were exposed to CoCl2 (350ppm) for 7 days, either alone, or in combination with COG (100 and 200mg/kg) or gallic acid (120mg/kg). CoCl2 given alone, caused significant increases (pgallic acid treatment significantly reduced (pgallic acid by modulation of CoCl2-induced alterations in blood pressure, antioxidant status and pro-apoptotic caspase 8 in Wistar rats.

  18. Molecular and acute temperature stress response characterizations of caspase-8 gene in two mussels, Mytilus coruscus and Mytilus galloprovincialis.

    Science.gov (United States)

    Zhang, Duo; Wang, Hong-Wei; Yao, Cui-Luan

    2014-01-01

    The caspase family represents aspartate-specific cysteine proteases that play key roles in initiation of apoptosis in various cells response to environmental stress. In this study, two caspase-8 cDNA sequences were cloned from two Mytilus mussels, Mytilus coruscus (Mccaspase-8) and Mytilus galloprovincialis (Mgcaspase-8), respectively. The full-length cDNA of Mccaspase-8 was 1884bp, including a 5'-terminal untranslated region (UTR) of 140bp, a 3'-terminal UTR of 238bp and an open reading frame (ORF) of 1506bp encoding a polypeptide of 501 amino acids. The 1775bp full-length Mg caspase-8 cDNA sequence contained an ORF of 1488bp encoding a polypeptide of 495 amino acid residues, a 5'-UTR of 51bp and a 3'-UTR of 236bp. Both the Mccaspase-8 and Mgcaspase-8 amino acid sequences contained two highly conservative death effector domains (DEDs) at N-terminal, the caspase family domains P20 and P10 and the caspase family cysteine active site 'KPKLFFIQACQG'. Phylogenetic analysis revealed that Mccaspase-8 and Mgcaspase-8 were clustered with the caspase-8 from other organisms, with the close relationship with caspase-8 from mollusk. Quantitative real-time reverse transcription PCR (qRT-PCR) analysis indicated that the predominant transcripts of Mccaspase-8 were in mantle and gonad tissue of M. coruscus and the high expression levels of Mgcaspase-8 were in digestive gland and gill tissue of M. galloprovincialis, respectively. The impacts of temperature stress on Mccaspase-8 and Mgcaspase-8 expressions were tested in gill tissue and hemocytes of both species. Our results showed that both Mccaspase-8 and Mgcaspase-8 transcripts and caspase-8 activity in gill tissue and hemocytes could be induced significantly after cold and heat stress (pgalloprovincialis.

  19. Caspase-8 acts as a molecular rheostat to limit RIPK1- and MyD88-mediated dendritic cell activation.

    Science.gov (United States)

    Cuda, Carla M; Misharin, Alexander V; Gierut, Angelica K; Saber, Rana; Haines, G Kenneth; Hutcheson, Jack; Hedrick, Stephen M; Mohan, Chandra; Budinger, G Scott; Stehlik, Christian; Perlman, Harris

    2014-06-15

    Caspase-8, an executioner enzyme in the death receptor pathway, was shown to initiate apoptosis and suppress necroptosis. In this study, we identify a novel, cell death-independent role for caspase-8 in dendritic cells (DCs): DC-specific expression of caspase-8 prevents the onset of systemic autoimmunity. Failure to express caspase-8 has no effect on the lifespan of DCs but instead leads to an enhanced intrinsic activation and, subsequently, more mature and autoreactive lymphocytes. Uncontrolled TLR activation in a RIPK1-dependent manner is responsible for the enhanced functionality of caspase-8-deficient DCs, because deletion of the TLR-signaling mediator, MyD88, ameliorates systemic autoimmunity induced by caspase-8 deficiency. Taken together, these data demonstrate that caspase-8 functions in a cell type-specific manner and acts uniquely in DCs to maintain tolerance.

  20. Protective Roles for Caspase-8 and cFLIP in Adult Homeostasis

    Directory of Open Access Journals (Sweden)

    Ricardo Weinlich

    2013-10-01

    Full Text Available Caspase-8 or cellular FLICE-like inhibitor protein (cFLIP deficiency leads to embryonic lethality in mice due to defects in endothelial tissues. Caspase-8−/− and receptor-interacting protein kinase-3 (RIPK3−/−, but not cFLIP−/− and RIPK3−/−, double-knockout animals develop normally, indicating that caspase-8 antagonizes the lethal effects of RIPK3 during development. Here, we show that the acute deletion of caspase-8 in the gut of adult mice induces enterocyte death, disruption of tissue homeostasis, and inflammation, resulting in sepsis and mortality. Likewise, acute deletion of caspase-8 in a focal region of the skin induces local keratinocyte death, tissue disruption, and inflammation. Strikingly, RIPK3 ablation rescues both phenotypes. However, acute loss of cFLIP in the skin produces a similar phenotype that is not rescued by RIPK3 ablation. TNF neutralization protects from either acute loss of caspase-8 or cFLIP. These results demonstrate that caspase-8-mediated suppression of RIPK3-induced death is required not only during development but also for adult homeostasis. Furthermore, RIPK3-dependent inflammation is dispensable for the skin phenotype.

  1. Novel HTS strategy identifies TRAIL-sensitizing compounds acting specifically through the caspase-8 apoptotic axis.

    Directory of Open Access Journals (Sweden)

    Darren Finlay

    Full Text Available Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL is potentially a very important therapeutic as it shows selectivity for inducing apoptosis in cancer cells whilst normal cells are refractory. TRAIL binding to its cognate receptors, Death Receptors-4 and -5, leads to recruitment of caspase-8 and classical activation of downstream effector caspases, leading to apoptosis. As with many drugs however, TRAIL's usefulness is limited by resistance, either innate or acquired. We describe here the development of a novel 384-well high-throughput screening (HTS strategy for identifying potential TRAIL-sensitizing agents that act solely in a caspase-8 dependent manner. By utilizing a TRAIL resistant cell line lacking caspase-8 (NB7 compared to the same cells reconstituted with the wild-type protein, or with a catalytically inactive point mutant of caspase-8, we are able to identify compounds that act specifically through the caspase-8 axis, rather than through general toxicity. In addition, false positive hits can easily be "weeded out" in this assay due to their activity in cells lacking caspase-8-inducible activity. Screening of the library of pharmacologically active compounds (LOPAC was performed as both proof-of-concept and to discover potential unknown TRAIL sensitizers whose mechanism is caspase-8 mediated. We identified known TRAIL sensitizers from the library and identified new compounds that appear to sensitize specifically through caspase-8. In sum, we demonstrate proof-of-concept and discovery of novel compounds with a screening strategy optimized for the detection of caspase-8 pathway-specific TRAIL sensitizers. This screen was performed in the 384-well format, but could easily be further miniaturized, allows easy identification of artifactual false positives, and is highly scalable to accommodate diverse libraries.

  2. MAPK p38 and JNK have opposing activities on TRAIL-induced apoptosis activation in NSCLC H460 cells that involves RIP1 and caspase-8 and is mediated by Mcl-1.

    Science.gov (United States)

    Azijli, Kaamar; Yuvaraj, Saravanan; van Roosmalen, Ingrid; Flach, Koen; Giovannetti, Elisa; Peters, Godefridus J; de Jong, Steven; Kruyt, Frank A E

    2013-07-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can induce both caspase-dependent apoptosis and kinase activation in tumor cells. Here, we examined the consequences and mechanisms of TRAIL-induced MAPKs p38 and JNK in non-small cell lung cancer (NSCLC) cells. In apoptosis sensitive H460 cells, these kinases were phosphorylated, but not in resistant A549 cells. Time course experiments in H460 cells showed that induction of p38 phosphorylation preceded that of JNK. To explore the function of these kinases in apoptosis activation by TRAIL, chemical inhibitors or siRNAs were employed to impair JNK or p38 functioning. JNK activation counteracted TRAIL-induced apoptosis whereas activation of p38 stimulated apoptosis. Notably, the serine/threonine kinase RIP1 was cleaved following TRAIL treatment, concomitant with detectable JNK phosphorylation. Further examination of the role of RIP1 by short hairpin (sh)RNA-dependent knockdown or inhibition by necrostatin-1 showed that p38 can be phosphorylated in both RIP1-dependent and -independent manner, whereas JNK phosphorylation occurred independent of RIP1. On the other hand JNK appeared to suppress RIP1 cleavage via an unknown mechanism. In addition, only the activation of JNK by TRAIL was caspase-8-dependent. Finally, we identified Mcl-1, a known substrate for p38 and JNK, as a downstream modulator of JNK or p38 activity. Collectively, our data suggest in a subset of NSCLC cells a model in which TRAIL-induced activation of p38 and JNK have counteracting effects on Mcl-1 expression leading to pro- or anti-apoptotic effects, respectively. Strategies aiming to stimulate p38 and inhibit JNK may have benefit for TRAIL-based therapies in NSCLC.

  3. RIP3 Inhibits Inflammatory Hepatocarcinogenesis but Promotes Cholestasis by Controlling Caspase-8- and JNK-Dependent Compensatory Cell Proliferation

    Directory of Open Access Journals (Sweden)

    Mihael Vucur

    2013-08-01

    Full Text Available For years, the term “apoptosis” was used synonymously with programmed cell death. However, it was recently discovered that receptor interacting protein 3 (RIP3-dependent “necroptosis” represents an alternative programmed cell death pathway activated in many inflamed tissues. Here, we show in a genetic model of chronic hepatic inflammation that activation of RIP3 limits immune responses and compensatory proliferation of liver parenchymal cells (LPC by inhibiting Caspase-8-dependent activation of Jun-(N-terminal kinase in LPC and nonparenchymal liver cells. In this way, RIP3 inhibits intrahepatic tumor growth and impedes the Caspase-8-dependent establishment of specific chromosomal aberrations that mediate resistance to tumor-necrosis-factor-induced apoptosis and underlie hepatocarcinogenesis. Moreover, RIP3 promotes the development of jaundice and cholestasis, because its activation suppresses compensatory proliferation of cholangiocytes and hepatic stem cells. These findings demonstrate a function of RIP3 in regulating carcinogenesis and cholestasis. Controlling RIP3 or Caspase-8 might represent a chemopreventive or therapeutic strategy against hepatocellular carcinoma and biliary disease.

  4. Independent Induction of Caspase-8 and cFLIP Expression during Colorectal Carcinogenesis in Sporadic and HNPCC Adenomas and Carcinomas

    Directory of Open Access Journals (Sweden)

    D. M. Heijink

    2007-01-01

    Full Text Available Background: TNF-Related Apoptosis Inducing Ligand (TRAIL is a promising agent for the induction of apoptosis in neoplastic tissues. Important determinants of TRAIL sensitivity are two intracellular proteins of the TRAIL pathway, caspase-8 and its anti-apoptotic competitor cellular Flice-Like Inhibitory Protein (cFLIP. Methods: The aim of this study was to investigate basic expression of caspase-8 and cFLIP in normal colorectal epithelium (n = 20, colorectal adenomas (n = 66 and colorectal carcinomas (n = 44 using immunohistochemistry performed on both sporadic and Hereditary Non-Polyposis Colorectal Cancer (HNPCC or Lynch syndrome-associated adenomas and carcinomas. Results: Expression of both caspase-8 and cFLIP was similar in cases with sporadic and hereditary origin. Expression of caspase-8 in colorectal adenomas and carcinomas was increased when compared to normal colon tissue (P = 0.02. Nuclear, paranuclear as well as cytoplasmic localizations of caspase-8 were detected. Immunohistochemistry revealed an upregulation of cFLIP in colorectal carcinomas in comparison to normal epithelium and colorectal adenomas (P < 0.001. A large variation in the caspase-8/cFLIP ratio was observed between the individual adenomas and carcinomas. Conclusion: Caspase-8 and cFLIP are upregulated during colorectal carcinogenesis. Upregulation of caspase-8 and/or downregulation of cFLIP may be interesting approaches to maximize TRAIL sensitivity in colorectal neoplasms.

  5. New insights into the regulation of innate immunity by caspase-8.

    Science.gov (United States)

    Sagulenko, Vitaliya; Lawlor, Kate E; Vince, James E

    2016-01-13

    Caspase-8 is required for extrinsic apoptosis, but is also central for preventing a pro-inflammatory receptor interacting protein kinase (RIPK) 3-mixed lineage kinase domain-like (MLKL)-dependent cell death pathway termed necroptosis. Despite these critical cellular functions, the impact of capase-8 deletion in the myeloid cell lineage, which forms the basis for innate immune responses, has remained unclear. In a recent article in Arthritis Research & Therapy, Cuda et al. report that myeloid cell-restricted caspase-8 loss leads to a very mild RIPK3-dependent inflammatory phenotype. The presented results suggest that inflammation does not arise exclusively because of RIPK3-mediated necroptotic death but that, in the absence of caspase-8, RIPK1 and RIPK3 enhance microbiome-driven Toll-like receptor-induced pro-inflammatory cytokine production.

  6. TRAF2 Sets a threshold for extrinsic apoptosis by tagging caspase-8 with a ubiquitin shutoff timer.

    Science.gov (United States)

    Gonzalvez, Francois; Lawrence, David; Yang, Becky; Yee, Sharon; Pitti, Robert; Marsters, Scot; Pham, Victoria C; Stephan, Jean-Philippe; Lill, Jennie; Ashkenazi, Avi

    2012-12-28

    Apoptotic caspase activation mechanisms are well defined, yet inactivation modes remain unclear. The death receptors (DRs), DR4, DR5, and Fas, transduce cell-extrinsic apoptotic signals by recruiting caspase-8 into a death-inducing signaling complex (DISC). At the DISC, Cullin3-dependent polyubiquitination on the small catalytic subunit of caspase-8 augments stimulation. Here we report that tumor necrosis factor receptor-associated factor 2 (TRAF2) interacts with caspase-8 at the DISC, downstream of Cullin3. TRAF2 directly mediates RING-dependent, K48-linked polyubiquitination on the large catalytic domain of caspase-8. This modification destines activated caspase-8 molecules to rapid proteasomal degradation upon autoprocessing and cytoplasmic translocation. TRAF2 depletion lowers the signal threshold for DR-mediated apoptosis, altering cell life versus death decisions in vitro and in vivo. Thus, TRAF2 sets a critical barrier for cell-extrinsic apoptosis commitment by tagging activated caspase-8 with a K48-ubiquitin shutoff timer. These results may have important implications for caspase regulation mechanisms.

  7. The human papillomavirus (HPV) E6 oncoproteins promotes nuclear localization of active caspase 8

    Energy Technology Data Exchange (ETDEWEB)

    Manzo-Merino, Joaquin [Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México. Av. San Fernando No. 22, Col. Sección XVI, Tlalpan 14080 (Mexico); Massimi, Paola [International Centre for Genetic Engineering and Biotechnology, Padriciano 99, I-34149 Trieste (Italy); Lizano, Marcela, E-mail: lizanosoberon@gmail.com [Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, México/Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México. Av. San Fernando No. 22, Col. Sección XVI, Tlalpan 14080 (Mexico); Banks, Lawrence, E-mail: banks@icgeb.org [International Centre for Genetic Engineering and Biotechnology, Padriciano 99, I-34149 Trieste (Italy)

    2014-02-15

    The HPV-16 E6 and E6{sup ⁎} proteins have been shown previously to be capable of regulating caspase 8 activity. We now show that the capacity of E6 to interact with caspase 8 is common to diverse HPV types, being also seen with HPV-11 E6, HPV-18 E6 and HPV-18 E6{sup ⁎}. Unlike most E6-interacting partners, caspase 8 does not appear to be a major proteasomal target of E6, but instead E6 appears able to stimulate caspase 8 activation, without affecting the overall apoptotic activity. This would appear to be mediated in part by the ability of the HPV E6 oncoproteins to recruit active caspase 8 to the nucleus. - Highlights: • Multiple HPV E6 oncoproteins interact with the caspase 8 DED domain. • HPV E6 stimulates activation of caspase 8. • HPV E6 promotes nuclear accumulation of caspase 8.

  8. Interaction between endoplasmic reticulum stress and caspase 8 activation in retrovirus MoMuLV-ts1-infected astrocytes.

    Science.gov (United States)

    Liu, Na; Scofield, Virginia L; Qiang, Wenan; Yan, Mingshan; Kuang, Xianghong; Wong, Paul K Y

    2006-05-10

    The murine retrovirus, MoMuLV-ts1, induces progressive paralysis and immune deficiency in FVB/N mice. We have reported previously that ts1 infection causes apoptosis in astrocytes via endoplasmic reticulum (ER) and mitochondrial stress (Liu, N., Kuang, X., Kim, H.T., Stoica, G., Qiang, W., Scofield, V.L., Wong, P.K.Y. Wong. 2004. Possible involvement of both endoplasmic reticulum- and mitochondria-dependent pathways in MoMuLV-ts1-induced apoptosis in astrocytes. J. NeuroVirol. 10, 189-198). In the present study, we show that caspase 8 activation in these cells is mediated through ER stress-associated elevation of death receptor DR5 and the C/EBP homologous protein (GADD153/CHOP), an ER stress-initiated transcription factor, rather than through TNFalpha and TNF-R1 interactions on the cell surface. Treatment with Z-IETD-FMK, a specific inhibitor of caspase 8 enzymatic activity, reduced ER stress by two mechanisms: by inhibiting caspase 8 activation, and by preventing cleavage of the ER-associated membrane protein BAP31 into BAP20, which exacerbates the ER stress response. These findings suggest that caspase 8- and ER stress-associated apoptotic pathways are linked in ts1-infected astrocytes.

  9. Caspase-8 as an Effector and Regulator of NLRP3 Inflammasome Signaling.

    Science.gov (United States)

    Antonopoulos, Christina; Russo, Hana M; El Sanadi, Caroline; Martin, Bradley N; Li, Xiaoxia; Kaiser, William J; Mocarski, Edward S; Dubyak, George R

    2015-08-14

    We recently described the induction of noncanonical IL-1β processing via caspase-8 recruited to ripoptosome signaling platforms in myeloid leukocytes. Here, we demonstrate that activated NLRP3·ASC inflammasomes recruit caspase-8 to drive IL-1β processing in murine bone marrow-derived dendritic cells (BMDC) independent of caspase-1 and -11. Sustained stimulation (>2 h) of LPS-primed caspase-1-deficient (Casp1/11(-/-)) BMDC with the canonical NLRP3 inflammasome agonist nigericin results in release of bioactive IL-1β in conjunction with robust caspase-8 activation. This IL-1β processing and caspase-8 activation do not proceed in Nlrp3(-/-) or Asc(-/-) BMDC and are suppressed by pharmacological inhibition of caspase-8, indicating that caspase-8 can act as a direct IL-1β-converting enzyme during NLRP3 inflammasome activation. In contrast to the rapid caspase-1-mediated death of wild type (WT) BMDC via NLRP3-dependent pyroptosis, nigericin-stimulated Casp1/11(-/-) BMDC exhibit markedly delayed cell death via NLRP3-dependent apoptosis. Biochemical analyses of WT and Casp1/11(-/-) BMDC indicated that caspase-8 is proteolytically processed within detergent-insoluble ASC-enriched protein complexes prior to extracellular export during nigericin treatment. Although nigericin-stimulated caspase-1 activation and activity are only modestly attenuated in caspase-8-deficient (Casp8(-/-)Rip3(-/-)) BMDC, these cells do not exhibit the rapid loss of viability of WT cells. These results support a contribution of caspase-8 to both IL-1β production and regulated death signaling via NLRP3 inflammasomes. In the absence of caspase-1, NLRP3 inflammasomes directly utilize caspase-8 as both a pro-apoptotic initiator and major IL-1β-converting protease. In the presence of caspase-1, caspase-8 acts as a positive modulator of the NLRP3-dependent caspase-1 signaling cascades that drive both IL-1β production and pyroptotic death.

  10. Contributions of caspase-8 and -9 to liver injury from CYP2E1-produced metabolites of halogenated hydrocarbons.

    Science.gov (United States)

    Ijiri, Yoshio; Kato, Ryuji; Sadamatsu, Maiko; Takano, Mina; Yasuda, Yuki; Tanaka, Fumiaki; Oishi, Chiyo; Imano, Hideki; Okada, Yoshikatsu; Tanaka, Kazuhiko; Hayashi, Tetsuya

    2017-01-12

    1. Drug-induced liver injury is difficult to predict at the pre-clinical stage. This study aimed to clarify the roles of caspase-8 and -9 in CYP2E1 metabolite-induced liver injury in both rats and cell cultures in vitro treated with carbon tetrachloride (CCl4), halothane or sevoflurane. The human hepatocarcinoma functional liver cell line was maintained in 3-dimensional culture alone or in co-culture with human acute monocytic leukemia cells. 2. In vivo, laboratory indices of liver dysfunction and histology were normal after administration of sevoflurane. CCl4 treatment increased blood AST/ALT levels, liver caspase-3 and -9 activities and liver malondialdehyde, accompanied by centrilobular hepatocyte necrosis. Halothane increased AST/ALT levels, caspase-3 and -8 activities (but not malondialdehyde) concomitant with widespread hepatotoxicity. In vitro, CCl4 treatment increased caspase-9 activity and decreased both mitochondrial membrane potential (MMP) and cell viability. In co-culture, halothane increased caspase-8 activity and decreased MMP and cellular viability. There were no toxic responses in CYP2E1 knockdown in monoculture and co-culture. 3. CYP2E1-inducing compounds play a pivotal role in halogenated hydrocarbon toxicity. 4. Changes in hepatocyte caspase-8 and -9 activities could be novel biomarkers of metabolites causing DILI, and in pre-clinical development of new pharmaceuticals can predict nascent DILI in the clinical stage.

  11. Caspase-8 Binding to Cardiolipin in Giant Unilamellar Vesicles Provides a Functional Docking Platform for Bid

    DEFF Research Database (Denmark)

    Jalmar, Olivier; Franc¸ois-Moutal, Liberty; García-Sáez, Ana-Jesus

    2013-01-01

    Caspase-8 is involved in death receptor-mediated apoptosis in type II cells, the proapoptotic programme of which is triggered by truncated Bid. Indeed, caspase-8 and Bid are the known intermediates of this signalling pathway. Cardiolipin has been shown to provide an anchor and an essential activa...

  12. Inhibition of caspase-8 activity reduces IFN-gamma expression by T cells from Leishmania major infection

    Directory of Open Access Journals (Sweden)

    Wânia F. Pereira

    2008-03-01

    Full Text Available Following infection with Leishmania major, T cell activation and apoptosis can be detected in draining lymph nodes of C57BL/6-infected mice. We investigated the mechanisms involved in apoptosis and cytokine expression following Tcellactivation. After two weeks of infection, apoptotic T cells were not detected in draining lymph nodes but activation with anti-CD3 induced apoptosis in both CD4 and CD8 T cells. Treatment with anti-FasLigand, caspase-8 or caspase- 9 inhibitors did not block activation-induced T-cell death. We also investigated whether the blockade of caspase-8 activity would affect the expression of type-1 or type-2 cytokines. At early stages of infection, both CD4 and CD8 T cells expressed IFN-gamma upon activation. Treatment with the caspase-8 inhibitor zIETD-fmk (benzyl-oxycarbonyl-Ile- Glu(OMe-Thr-Asp(OMe-fluoromethyl ketone reduced the proportion of CD8 T cells and IFN-gamma expression in both CD4 and CD8T cells. We conclude that a non apoptotic role of caspase-8 activity may be required for T cell-mediated type-1 responses during L. major infection.A ativação e a morte por apoptose de linfócitos T foram observadas em linfonodos drenantes de camundongos C57BL/6 infectados com Leishmania major. Investigamos os mecanismos envolvidos na apoptose e na expressão de citocinas após a ativação de linfócitos T. Após duas semanas de infecção, embora as células apoptóticas ainda não sejam detectadas em linfonodos drenantes, células T CD4 e CD8 sofrem apoptose após ativação com anti-CD3. O tratamento com anticorpo antagonista anti-Ligante de Fas, ou com inibidores das caspases-8 e 9, não bloqueou a morte induzida por ativação das células T. Investigamos também se a inibição da atividade da caspase-8 poderia afetar a expressão de citocinas tipo-1 ou tipo-2. Nos estágios iniciais da infecção, células T CD4 e CD8 de animais infectados com L. major expressaram IFN-gama após ativação. O tratamento com o inibidor

  13. Requirement for caspase-8 in NF-kappaB activation by antigen receptor.

    Science.gov (United States)

    Su, Helen; Bidère, Nicolas; Zheng, Lixin; Cubre, Alan; Sakai, Keiko; Dale, Janet; Salmena, Leonardo; Hakem, Razqallah; Straus, Stephen; Lenardo, Michael

    2005-03-04

    Caspase-8, a proapoptotic protease, has an essential role in lymphocyte activation and protective immunity. We show that caspase-8 deficiency (CED) in humans and mice specifically abolishes activation of the transcription factor nuclear factor kappaB (NF-kappaB) after stimulation through antigen receptors, Fc receptors, or Toll-like receptor 4 in T, B, and natural killer cells. Caspase-8 also causes the alphabeta complex of the inhibitor of NF-kappaB kinase (IKK) to associate with the upstream Bcl10-MALT1 (mucosa-associated lymphatic tissue) adapter complex. Recruitment of the IKKalpha, beta complex, its activation, and the nuclear translocation of NF-kappaB require enzyme activity of full-length caspase-8. These findings thus explain the paradoxical association of defective apoptosis and combined immunodeficiency in human CED.

  14. Hepatocyte caspase-8 is an essential modulator of steatohepatitis in rodents

    NARCIS (Netherlands)

    Hatting, M.; Zhao, G.; Schumacher, F.; Sellge, G.; Masaoudi, Al M.; Gaßler, N.; Boekschoten, M.V.; Müller, M.R.; Liedtke, C.; Cubero, F.J.; Trautwein, C.

    2013-01-01

    In human and murine models of nonalcoholic steatohepatitis (NASH), increased hepatocyte apoptosis is a critical mechanism contributing to inflammation and fibrogenesis. Caspase 8 (Casp8) is essential for death-receptor-mediated apoptosis activity and therefore its modulation might be critical for th

  15. The apoptotic initiator caspase-8: its functional ubiquity and genetic diversity during animal evolution.

    Science.gov (United States)

    Sakamaki, Kazuhiro; Shimizu, Kouhei; Iwata, Hiroaki; Imai, Kenichiro; Satou, Yutaka; Funayama, Noriko; Nozaki, Masami; Yajima, Mamiko; Nishimura, Osamu; Higuchi, Mayura; Chiba, Kumiko; Yoshimoto, Michi; Kimura, Haruna; Gracey, Andrew Y; Shimizu, Takashi; Tomii, Kentaro; Gotoh, Osamu; Akasaka, Koji; Sawasaki, Tatsuya; Miller, David J

    2014-12-01

    The caspases, a family of cysteine proteases, play multiple roles in apoptosis, inflammation, and cellular differentiation. Caspase-8 (Casp8), which was first identified in humans, functions as an initiator caspase in the apoptotic signaling mediated by cell-surface death receptors. To understand the evolution of function in the Casp8 protein family, casp8 orthologs were identified from a comprehensive range of vertebrates and invertebrates, including sponges and cnidarians, and characterized at both the gene and protein levels. Some introns have been conserved from cnidarians to mammals, but both losses and gains have also occurred; a new intron arose during teleost evolution, whereas in the ascidian Ciona intestinalis, the casp8 gene is intronless and is organized in an operon with a neighboring gene. Casp8 activities are near ubiquitous throughout the animal kingdom. Exogenous expression of a representative range of nonmammalian Casp8 proteins in cultured mammalian cells induced cell death, implying that these proteins possess proapoptotic activity. The cnidarian Casp8 proteins differ considerably from their bilaterian counterparts in terms of amino acid residues in the catalytic pocket, but display the same substrate specificity as human CASP8, highlighting the complexity of spatial structural interactions involved in enzymatic activity. Finally, it was confirmed that the interaction with an adaptor molecule, Fas-associated death domain protein, is also evolutionarily ancient. Thus, despite structural diversity and cooption to a variety of new functions, the ancient origins and near ubiquitous distribution of this activity across the animal kingdom emphasize the importance and utility of Casp8 as a central component of the metazoan molecular toolkit.

  16. Pharmacological modulation of caspase-8 in thymus-related medical conditions.

    Science.gov (United States)

    Pozzesi, Nicola; Fierabracci, Alessandra; Thuy, Trinh Thy; Martelli, Maria Paola; Liberati, Anna Marina; Ayroldi, Emira; Riccardi, Carlo; Delfino, Domenico V

    2014-10-01

    The thymus is a lymphoid organ that governs the development of a diverse T-cell repertoire capable of defending against nonself-antigens and avoiding autoimmunity. However, the thymus can also succumb to different diseases. Hypertrophic diseases, such as thymomas, are typically associated with impairment of negative selection, which leads to autoimmune disease, or disruption of positive selection, which results in immunodeficiency. Hypotrophic diseases of the thymus can manifest during acute infections, cancer, allogeneic bone marrow transplantation, or with aging. This condition leads to decreased immune function and can be treated by either replacing lost thymic tissue or by preventing thymic tissue death. Studies have demonstrated the critical role of caspase-8 in regulating apoptosis in the thymus. In this review, we discuss how pharmacological activation and inhibition of caspase-8 can be used to treat hypertrophic and hypotrophic diseases of the thymus, respectively, to improve its function.

  17. Effects of methylation status of caspase-8 promoter on antitumor activity of TRAIL to human gastric cancer cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Ru-gang; FANG Dian-chun; YANG Liu-qin; LUO Yuan-gang

    2004-01-01

    Objective: To study the effects of the methylation status of caspase-8 promoter on the antitumor activity of TRAIL to the human gastric cancer cells. Methods: The methylation of caspase-8 was measured with methylation specific PCR (MSP) and the antitomor capability of TRAIL to human gastric cancer cells was determined with MTT. Results: No methylation of caspase-8 in the human gastric cancer cells was found. The sensitivity of 5 lines of gastric cancer cells to the antitumor activity of TRAIL was different. The administration of the demethylation agent 5-Aza-2'-deoxycytidine ( 5-AzaCdR) increased the sensitivity of gastric cancer cells to TRAIL but did not change the methylation status of caspase-8 promoter in gastric cancer cells. Conclusion: 5-Aza-CdR increases the sensitivity of most of gastric cancer cells to TRAIL but caspase-8 is not involved in the antitumor activity of TRAIL.

  18. Maternal hyperglycemia activates an ASK1-FoxO3a-caspase 8 pathway that leads to embryonic neural tube defects.

    Science.gov (United States)

    Yang, Peixin; Li, Xuezheng; Xu, Cheng; Eckert, Richard L; Reece, E Albert; Zielke, Horst Ronald; Wang, Fang

    2013-08-27

    Neural tube defects result from failure to completely close neural tubes during development. Maternal diabetes is a substantial risk factor for neural tube defects, and available evidence suggests that the mechanism that links hyperglycemia to neural tube defects involves oxidative stress and apoptosis. We demonstrated that maternal hyperglycemia correlated with activation of the apoptosis signal-regulating kinase 1 (ASK1) in the developing neural tube, and Ask1 gene deletion was associated with reduced neuroepithelial cell apoptosis and development of neural tube defects. ASK1 activation stimulated the activity of the transcription factor FoxO3a, which increased the abundance of the apoptosis-promoting adaptor protein TRADD, leading to activation of caspase 8. Hyperglycemia-induced apoptosis and the development of neural tube defects were reduced with genetic ablation of either FoxO3a or Casp8 or inhibition of ASK1 by thioredoxin. Examination of human neural tissues affected by neural tube defects revealed increased activation or abundance of ASK1, FoxO3a, TRADD, and caspase 8. Thus, activation of an ASK1-FoxO3a-TRADD-caspase 8 pathway participates in the development of neural tube defects, which could be prevented by inhibiting intermediates in this cascade.

  19. Expression and prognostic significance of APAF-1, caspase-8 and caspase-9 in stage II/III colon carcinoma: caspase-8 and caspase-9 is associated with poor prognosis.

    Science.gov (United States)

    Sträter, Jörn; Herter, Ines; Merkel, Gaby; Hinz, Ulf; Weitz, Jürgen; Möller, Peter

    2010-08-15

    Apoptosis protease activating factor-1 (APAF-1), caspase-8 and caspase-9 are important factors in the execution of death signals. To study their prognostic influence in colon carcinoma, expression of APAF-1, caspase-8 and caspase-9 was determined by immunohistochemistry in normal colon mucosa (n = 8) and R0-resected stage II/III colon carcinomas (n >or= 124) using a semiquantitative score. Staining results were correlated with disease-free survival by Kaplan-Meier estimates, and multivariate Cox analyses were performed. In normal colon, APAF-1 and caspase-8 are most strongly expressed in the luminal surface epithelium, whereas caspase-9 is expressed all along the crypt axis. In colon carcinomas, there is considerable variability in the expression of these proapoptotic factors, although complete loss of caspase-8 and caspase-9 is rare. APAF-1 expression did not correlate with disease-free survival. Instead, both expression of caspase-9 and high-level expression of caspase-8 in a majority of tumor cells were significantly associated with adverse prognosis (p = 0.004 and p = 0.029, respectively). The influence of caspase-8 expression was mainly seen in patients with stage III colon carcinoma (p = 0.011), whereas the prognostic influence of caspase-9 expression was significant in stage II cases (p = 0.037) and just failed to be significant in stage III tumors (p = 0.0581). After adjusting for confounding factors in a multivariate Cox analysis, the effect of caspase-9 in predicting disease-free survival was confirmed (p = 0.003). Our data suggest that, in colon carcinomas, expression of caspase-8 and caspase-9 is significantly associated with poor survival. Caspase-9 may be an independent prognosticator in colon carcinoma.

  20. Deletion of caspase-8 in mouse myeloid cells blocks microglia pro-inflammatory activation and confers protection in MPTP neurodegeneration model.

    Science.gov (United States)

    Kavanagh, Edel; Burguillos, Miguel Angel; Carrillo-Jimenez, Alejandro; Oliva-Martin, María José; Santiago, Martiniano; Rodhe, Johanna; Joseph, Bertrand; Venero, Jose Luis

    2015-09-01

    Increasing evidence involves sustained pro-inflammatory microglia activation in the pathogenesis of different neurodegenerative diseases, particularly Parkinson's disease (PD). We recently uncovered a completely novel and unexpected role for caspase-8 and its downstream substrates caspase-3/7 in the control of microglia activation and associated neurotoxicity to dopaminergic cells. To demonstrate the genetic evidence, mice bearing a floxed allele ofCASP8 were crossed onto a transgenic line expressing Cre under the control of Lysozyme 2 gene. Analysis of caspase-8 gene deletion in brain microglia demonstrated a high efficiency in activated but not in resident microglia. Mice were challenged with lipopolysaccharide, a potent inducer of microglia activation, or with MPTP, which promotes specific dopaminergic cell damage and consequent reactive microgliosis. In neither of these models, CASP8 deletion appeared to affect the overall number of microglia expressing the pan specific microglia marker, Iba1. In contrast, CD16/CD32 expression, a microglial pro-inflammatory marker, was found to be negatively affected upon CASP8 deletion. Expression of additional proinflammatory markers were also found to be reduced in response to lipopolysaccharide. Of importance, reduced pro-inflammatory microglia activation was accompanied by a significant protection of the nigro-striatal dopaminergic system in the MPTP mouse model of PD.

  1. TNF-α has tropic rather than apoptotic activity in human hematopoietic progenitors: involvement of TNF receptor-1 and caspase-8.

    Science.gov (United States)

    Mizrahi, Keren; Stein, Jerry; Yaniv, Isaac; Kaplan, Offer; Askenasy, Nadir

    2013-01-01

    Tumor necrosis factor-α (TNF-α) has been suggested to exert detrimental effects on hematopoietic progenitor function that might limit the success of transplants. In this study, we assessed the influences of TNF-α and its two cognate receptors on the function of fresh umbilical cord blood (UCB) and cryopreserved mobilized peripheral blood (mPB). CD34(+) progenitors from both sources are less susceptible to spontaneous apoptosis than lineage-committed cells and are not induced into apoptosis by TNF-α. Consequently, the activity of UCB-derived severe combined immune deficiency (SCID) reconstituting cells and long-term culture-initiating cells is unaffected by this cytokine. On the contrary, transient exposure of cells from both sources to TNF-α stimulates the activity of myeloid progenitors, which persists in vivo in UCB cell transplants. Progenitor stimulation is selectively mediated by TNF-R1 and involves activation of caspase-8, without redundant activity of TNF-R2. Despite significant differences between fresh UCB cells and cryopreserved mPB cells in susceptibility to apoptosis and time to activation, TNF-α is primarily involved in tropic signaling in hematopoietic progenitors from both sources. Cytokine-mediated tropism cautions against TNF-α neutralization under conditions of stress hematopoiesis and may be particularly beneficial in overcoming the limitations of UCB cell transplants.

  2. TRAIL Activates a Caspase 9/7-Dependent Pathway in Caspase 8/10-Defective SK-N-SH Neuroblastoma Cells with Two Functional End Points: Induction of Apoptosis and PGE2 Release

    Directory of Open Access Journals (Sweden)

    Giorgio Zauli

    2003-09-01

    Full Text Available Most neuroblastoma cell lines do not express apical caspases 8 and 10, which play a key role in mediating tumor necrosis factor-related apoptosis-inducing ligand (TRAIL cytotoxicity in a variety of malignant cell types. In this study, we demonstrated that TRAIL induced a moderate but significant increase of apoptosis in the caspase 8/10-deficient SK-N-SH neuroblastoma cell line, through activation of a novel caspase 9/7 pathway. Concomitant to the induction of apoptosis, TRAIL also promoted a significant increase of prostaglandin E2 (PGE2 release by SKN-SH cells. Moreover, coadministration of TRAIL plus indomethacin, a pharmacological inhibitor of cyclooxygenase (COX, showed an additive effect on SKN-SH cell death. In spite of the ability of TRAIL to promote the phosphorylation of both ERKi/2 and p38/MAPK, which have been involved in the control of COX expression/activity, neither PD98059 nor SB203580, pharmacological inhibitors of the ERKi/2 and p38/MAPK pathways, respectively, affected either PGE2 production or apoptosis induced by TRAIL. Finally, both induction of apoptosis and PGE2 release were completely abrogated by the broad caspase inhibitor z-VAD4mk, suggesting that both biologic end points were regulated in SK-N-SH cells through a caspase 9/7-dependent pathway.

  3. Dietary n-3 PUFAs augment caspase 8 activation in Staphylococcal aureus enterotoxin B stimulated T-cells.

    Science.gov (United States)

    Gill, R; Jen, K L; McCabe, M J J; Rosenspire, A

    2016-10-15

    Epidemiological studies have linked consumption of n-3 PUFAs with a variety of beneficial health benefits, particularly with respect to putative anti-inflammatory effects. Unfortunately, many of these results remain somewhat controversial because in most instances there has not been a linkage to specific molecular mechanisms. For instance, dietary exposure to low levels of mercury has been shown to be damaging to neural development, but concomitant ingestion of n-3 PUFAs as occurs during consumption of fish, has been shown to counteract the detrimental effects. As the mechanisms mediating the neurotoxicity of environmental mercury are not fully delineated, it is difficult to conceptualize a testable molecular mechanism explaining how n-3 PUFAs negate its neurotoxic effects. However, environmental exposure to mercury also has been linked to increased autoimmunity. By way of a molecular understanding of this immuno-toxic association, disruption of CD95 signaling is well established as a triggering factor for autoimmunity, and we have previously shown that environmentally relevant in vitro and dietary exposures to mercury interfere with CD95 signaling. In particular we have shown that activation of caspase 8, as well as downstream activation of caspase 3, in response to CD95 agonist stimulation is depressed by mercury. More recently we have shown in vitro that the n-3 PUFA docosahexaenoic acid counteracts the negative effect of mercury on CD95 signaling by restoring caspase activity. We hypothesized that concomitant ingestion of n-3 PUFAs with mercury might be protective from the immuno-toxic effects of mercury, as it is with mercury's neuro-toxic effects, and in the case of immuno-toxicity this would be related to restoration of CD95 signal strength. We now show that dietary ingestion of n-3 PUFAs generally promotes CD95 signaling by upregulating caspase 8 activation. Apart from accounting for the ability of n-3 PUFAs to specifically counteract autoimmune sequelae of

  4. Synergistic Effect of Subtoxic-dose Cisplatin and TRAIL to Mediate Apoptosis by Down-regulating Decoy Receptor 2 and Up-regulating Caspase-8, Caspase-9 and Bax Expression on NCI-H460 and A549 Cells

    Directory of Open Access Journals (Sweden)

    Xiaoyan Zhang

    2013-05-01

    Full Text Available Objective(s: Although tumor necrosis factor-related apoptosis-inducing ligand (TRAIL can selectively induce apoptosis in tumor cells, more than half of tumors including non-small cell lung cancer (NSCLC exhibit TRAIL-resistance. The purpose of this study was to determine whether subtoxic-dose cisplatin and TRAIL could synergistically enhance apoptosis on NSCLC cells and investigate its underlying mechanisms. Materials and Methods:NCI-H460 and A549 cells were treated with TRAIL alone, cisplatin alone or combination treatment in this study. The cytotoxicity was evaluated according to Sulforhodamine B assay, and apoptosis was examined using Hoechst 33342 staining and flow cytometry. The mRNA and protein levels of TRAIL receptors and apoptotic proteins including caspase-8, caspase-9, Bcl-2 and Bax were determined by RT-PCR and Western blotting, respectively. Results:Our results showed that NCI-H460 cells were sensitive to TRAIL, whereas A549 cells were resistant. However, subtoxic-dose cisplatin could enhance the both cells to TRAIL-mediated cell proliferation inhibition and apoptosis. The underlying mechanisms might be associated with the down-regulation of DcR2 and up-regulation of Caspase-8, Caspase-9 and Bax. Conclusion:Subtoxic-dose cisplatin could enhance both TRAIL- sensitive and TRAIL- resistant NSCLC cells to TRAIL-mediated apoptosis. These findings motivated further studies to evaluate such a combinatory therapeutic strategy against NSCLC in the animal models.

  5. Synergistic effect of fisetin combined with sorafenib in human cervical cancer HeLa cells through activation of death receptor-5 mediated caspase-8/caspase-3 and the mitochondria-dependent apoptotic pathway.

    Science.gov (United States)

    Lin, Ming-Te; Lin, Chia-Liang; Lin, Tzu-Yu; Cheng, Chun-Wen; Yang, Shun-Fa; Lin, Chu-Liang; Wu, Chih-Chien; Hsieh, Yi-Hsien; Tsai, Jen-Pi

    2016-05-01

    Combining antitumor agents with bioactive compounds is a potential strategy for improving the effect of chemotherapy on cancer cells. The goal of this study was to elucidate the antitumor effect of the flavonoid, fisetin, combined with the multikinase inhibitor, sorafenib, against human cervical cancer cells in vitro and in vivo. The combination of fisetin and sorafenib synergistically induced apoptosis in HeLa cells, which is accompanied by a marked increase in loss of mitochondrial membrane potential. Apoptosis induction was achieved by caspase-3 and caspase-8 activation which increased the ratio of Bax/Bcl-2 and caused the subsequent cleavage of PARP level while disrupting the mitochondrial membrane potential in HeLa cells. Decreased Bax/Bcl-2 ratio level and mitochondrial membrane potential were also observed in siDR5-treated HeLa cells. In addition, in vivo studies revealed that the combined fisetin and sorafenib treatment was clearly superior to sorafenib treatment alone using a HeLa xenograft model. Our study showed that the combination of fisetin and sorafenib exerted better synergistic effects in vitro and in vivo than either agent used alone against human cervical cancer, and this synergism was based on apoptotic potential through a mitochondrial- and DR5-dependent caspase-8/caspase-3 signaling pathway. This combined fisetin and sorafenib treatment represents a novel therapeutic strategy for further clinical developments in advanced cervical cancer.

  6. Update of the Caspase-3 and Caspase-8 in lichen planus%Caspase-3和 Caspase-8与扁平苔藓的研究进展

    Institute of Scientific and Technical Information of China (English)

    李帅; 栗玉珍

    2016-01-01

    Caspase-3 and Caspase-8 are important proteins in the process of apoptosis. Caspase-8 can start apoptotic reaction which can mediate apoptosis in a variety of ways. Caspase-3 can accelerate cell apop-tosis after activation. The new studies reported high expression of Caspase-3, Caspase-8 in lichen planus le-sions, especially in the basal layer which showed that Caspase-3, Caspase-8 might be involved in the apop-totic response process of lichen planus.%Caspase-3和 Caspase-8作为含半胱氨酸的天冬氨酸蛋白水解酶家族成员,是参与细胞凋亡过程的两个重要蛋白。 Caspase-8是重要的凋亡启动蛋白,可通过多种途径介导凋亡反应的发生, Caspase-3作为凋亡过程中的执行者,活化后可以加速细胞凋亡的发生。最新研究发现 Caspase-3和Caspase-8在扁平苔藓皮损处(特别是基底层处)高表达,表明两者可能参与了扁平苔藓的凋亡过程,并导致了本病的病理变化。

  7. A delay prior to mitotic entry triggers caspase 8-dependent cell death in p53-deficient Hela and HCT-116 cells.

    Science.gov (United States)

    Silva, Victoria C; Plooster, Melissa; Leung, Jessica C; Cassimeris, Lynne

    2015-01-01

    Stathmin/Oncoprotein 18, a microtubule destabilizing protein, is required for survival of p53-deficient cells. Stathmin-depleted cells are slower to enter mitosis, but whether delayed mitotic entry triggers cell death or whether stathmin has a separate pro-survival function was unknown. To test these possibilities, we abrogated the cell cycle delay by inhibiting Wee1 in synchronized, stathmin-depleted cells and found that apoptosis was reduced to control levels. Synchronized cells treated with a 4 hour pulse of inhibitors to CDK1 or both Aurora A and PLK1 delayed mitotic entry and apoptosis was triggered only in p53-deficient cells. We did not detect mitotic defects downstream of the delayed mitotic entry, indicating that cell death is activated by a mechanism distinct from those activated by prolonged mitotic arrest. Cell death is triggered by initiator caspase 8, based on its cleavage to the active form and by rescue of viability after caspase 8 depletion or treatment with a caspase 8 inhibitor. In contrast, initiator caspase 9, activated by prolonged mitotic arrest, is not activated and is not required for apoptosis under our experimental conditions. P53 upregulates expression of cFLIPL, a protein that blocks caspase 8 activation. cFLIPL levels are lower in cells lacking p53 and these levels are reduced to a greater extent after stathmin depletion. Expression of FLAG-tagged cFLIPL in p53-deficient cells rescues them from apoptosis triggered by stathmin depletion or CDK1 inhibition during G2. These data indicate that a cell cycle delay in G2 activates caspase 8 to initiate apoptosis specifically in p53-deficient cells.

  8. Caspase 8 and maspin are downregulated in breast cancer cells due to CpG site promoter methylation

    Directory of Open Access Journals (Sweden)

    Koeffler Phillip

    2010-02-01

    Full Text Available Abstract Background Epigenetic changes associated with promoter DNA methylation results in silencing of several tumor suppressor genes that lead to increased risk for tumor formation and for progression of the cancer. Methods Methylation specific PCR (MSP and bisulfite sequencing were used for determination of proapoptotic gene Caspase 8 (CASP8 and the tumor suppressor gene maspin promoter methylation in four breast cancer and two non-tumorigenic breast cell lines. Involvement of histone H3 methylation in those cell lines were examined by CHIP assay. Results The CpG sites in the promoter region of CASP8 and maspin were methylated in all four breast cancer cell lines but not in two non-tumorigenic breast cell lines. Demethylation agent 5-aza-2'-deoxycytidine (5-aza-dc selectively inhibits DNA methyltransferases, DNMT3a and DNMT3b, and restored CASP8 and maspin gene expression in breast cancer cells. 5-aza-dc also reduced histone H3k9me2 occupancy on CASP8 promoter in SKBR3cells, but not in MCF-7 cells. Combination of histone deacetylase inhibitor Trichostatin A (TSA and 5-aza-dc significant decrease in nuclear expression of Di-methyl histone H3-Lys27 and slight increase in acetyl histone H3-Lys9 in MCF-7 cells. CASP8 mRNA and protein level in MCF-7 cells were increased by the 5-aza-dc in combination with TSA. Data from our study also demonstrated that treatment with 5-FU caused a significant increase in unmethylated CASP8 and in CASP8 mRNA in all 3 cancer lines. Conclusions CASP8 and maspin expression were reduced in breast cancer cells due to promoter methylation. Selective application of demethylating agents could offer novel therapeutic opportunities in breast cancer.

  9. BmDredd is an initiator caspase and participates in Emodin-induced apoptosis in the silkworm, Bombyx mori.

    Science.gov (United States)

    Wang, La; Song, Juan; Bao, Xi-Yan; Chen, Peng; Yi, Hua-Shan; Pan, Min-Hui; Lu, Cheng

    2016-10-15

    The identification and analysis of the caspases is essential to research into apoptosis in lepidoptera insects. The domesticated silkworm, Bombyx mori, is the model system for lepidopterans. In this study, we cloned and characterized a B. mori Dredd gene, BmDredd, the proposed insect homologue of human caspase-8, which encoded a polypeptide of 543 amino acids. BmDredd possesses a long N-terminal prodomain, a p20 domain, and a p10 domain. When transiently expressed in Escherichia coli cells, BmDredd underwent spontaneous cleavage and exhibited high proteolytic activity for caspase-8 substrate but relatively low for caspase-3 or -9 substrate. In addition, BmDredd induced apoptosis when transiently expressed in BmN-SWU1 cells, an ovarian cell line of B. mori. Moreover, after the treatment of Emodin, a novel apoptosis inducer, endogenous BmDredd expression level, the caspase-8 activity and the apoptotic rate increased notably in BmN-SWU1 cells. When BmDredd was subjected to interference in BmN-SWU1 cells and Emodin treatment, BmDredd expression levels decreased and the apoptotic rate also decreased significantly. These results suggest BmDredd is the homologue of human caspase-8 and plays a role in Emodin-induced apoptosis in BmN-SWU1 cells of B. mori.

  10. 微小RNA-1187对半胱天冬氨酸蛋白酶-8介导的肝细胞凋亡调控的作用%Regulatory role of microRNA-1187 in caspase-8 mediated hepatocyte apoptosis

    Institute of Scientific and Technical Information of China (English)

    余东山; 安方梅; 龚邦东; 赵钢德; 项晓刚; 林兰意; 俞红; 王晖; 谢青

    2011-01-01

    Objective To observe the regulatory role of microRNA-1187(miR-1187)in hepatocyte apoptosis through miR-1187 targeting regulation of caspase-8 mRNA expression.Methods The acute liver failure model was established by injection of D-galactosamine plus lipopolysaccharides(LPS)in BALB/c mice.The liver tissues were collected for LNA-miRNA array analysis and functional analysis of genes targeted by miRNA.Embryonic murine hepatocyte cell line 2(BNL-CL2)was cultivated in vitro and treated with tumor necrosis factor(TNF)-α and D-galactosamine to induce the transfection of miR-1187 in transfected group or untransfected group.The expressions of miR-1187 and caspase-8 mRNA were detected by real-time polymeramse chain reaction(PCR)and caspase-8 protein was determined by Western blot.The apoptosis rate was detected by flow cytometry.The comparison of means between groups was done by t test.Results The miR-1187 signal was deceased with the development of acute liver failure.The 3'UTR of caspase-8 mRNA had direct binding sites with miR1187.In BNL-CL2 cell experiments,miR-1187 was down-regulated in untransfected group and decreased more slowly in transfected group(t=6.371,P<0.01).The expression of caspase-8 mRNA was up-regulated in untransfected group and increased less in transfected group(t=4.539,P<0.01).The apoptosis rate in transfected group was significantly lower than untransfected group(t=3.365,P<0.05).Conclusios miR-1187 is one of inhibitors of hepatocyte apoptosis.High expression of miR-1187 could regulate the expression of caspase-8 mRNA,thus inhibit the apoptosis of hepatocytes.%目的 通过微小RNA-1187(miR-1187)靶向调节半胱天冬氨酸蛋白酶-8(caspase-8)mRNA的表达,观察miR-1187对肝细胞凋亡的调控作用.方法 应用D-氨基半乳糖联合脂多糖构建BALB/c小鼠急性肝功能衰竭模型,取肝组织进行锁核酸(LNA)-miRNA微阵列分析和miRNA靶基因功能分析.体外培养胚胎小鼠肝细胞株2(BNL-CL2细胞),予以TNF

  11. Mycobacterium tuberculosis-infected human monocytes down-regulate microglial MMP-2 secretion in CNS tuberculosis via TNFα, NFκB, p38 and caspase 8 dependent pathways

    Directory of Open Access Journals (Sweden)

    Elkington Paul T

    2011-05-01

    Full Text Available Abstract Tuberculosis (TB of the central nervous system (CNS is a deadly disease characterized by extensive tissue destruction, driven by molecules such as Matrix Metalloproteinase-2 (MMP-2 which targets CNS-specific substrates. In a simplified cellular model of CNS TB, we demonstrated that conditioned medium from Mycobacterium tuberculosis-infected primary human monocytes (CoMTb, but not direct infection, unexpectedly down-regulates constitutive microglial MMP-2 gene expression and secretion by 72.8% at 24 hours, sustained up to 96 hours (P M.tb-infected monocyte-dependent networks paradoxically involves the pro-inflammatory mediators TNF-α, p38 MAP kinase and NFκB in addition to a novel caspase 8-dependent pathway.

  12. Galanin Protects from Caspase-8/12-initiated Neuronal Apoptosis in the Ischemic Mouse Brain via GalR1

    Science.gov (United States)

    Li, Yun; Mei, Zhu; Liu, Shuiqiao; Wang, Tong; Li, Hui; Li, Xiao-Xiao; Han, Song; Yang, Yutao; Li, Junfa; Xu, Zhi-Qing David

    2017-01-01

    Galanin (GAL) plays key role in many pathophysiological processes, but its role in ischemic stroke remains unclear. Here, the models of 1 h middle cerebral artery occlusion (MCAO)/1-7 d reperfusion (R)-induced ischemic stroke and in vitro cell ischemia of 1 h oxygen-glucose deprivation (OGD)/24 h reoxygenation in primary cultured cortical neurons were used to explore GAL’s effects and its underlying mechanisms. The results showed significant increases of GAL protein levels in the peri-infarct region (P) and infarct core (I) within 48 h R of MCAO mice (p<0.001). The RT-qPCR results also demonstrated significant increases of GAL mRNA during 24-48 h R (p<0.001), and GAL receptors GalR1-2 (but not 3) mRNA levels in the P region at 24 h R of MCAO mice (p<0.001). Furthermore, the significant decrease of infarct volume (p<0.05) and improved neurological outcome (p<0.001-0.05) were observed in MCAO mice following 1 h pre- or 6 h post-treatment of GAL during 1-7 d reperfusion. GalR1 was confirmed as the receptor responsible for GAL-induced neuroprotection by using GalR2/3 agonist AR-M1896 and Lentivirus-based RNAi knockdown of GalR1. GAL treatment inhibited Caspase-3 activation through the upstream initiators Capsases-8/-12 (not Caspase-9) in both P region and OGD-treated cortical neurons. Meanwhile, GAL’s neuroprotective effect was not observed in cortical neurons from conventional protein kinase C (cPKC) γ knockout mice. These results suggested that exogenous GAL protects the brain from ischemic injury by inhibiting Capsase-8/12-initiated apoptosis, possibly mediated by GalR1 via the cPKCγ signaling pathway. PMID:28203483

  13. Caspase-8、NF-kB在经Gen处理后的大鼠子宫内膜异位症中的表达%Expressions of Caspase-8 and NF-kB in endometriosis of rats treated with genistein

    Institute of Scientific and Technical Information of China (English)

    崔慧星; 彭小星; 文兰英; 金延泽

    2014-01-01

    目的:观察Caspase-8,NF-kB在经金雀异黄素(Genistein,Gen)处理后的大鼠子宫内膜异位症中的表达,初步探讨金雀异黄素对大鼠子宫内膜异位症治疗的作用机制.方法:采用外科手术法建立大鼠EMs模型,将大鼠随机分为Ⅰ组15只(对照组)、Ⅱ组15只(Gen低剂量治疗组,0.5 mg·kg-1·d-1)、Ⅲ组15只(Gen高剂量治疗组,50mg·kg-1 ·d-1).连续皮下注射药物12周,再次检测移植物体积变化,运用HE染色观察移植内膜组织的病理改变,免疫组化法检测大鼠异位内膜组织中Caspase-8及NF-kB的表达.结果:移植物外观:与对照组比较,Gen高剂量治疗组移植物体积明显缩小(P<0.01).病理学变化:与对照组比较,Gen治疗组移植腺体可见部分萎缩、变矮、消失,其上皮细胞层厚度变薄,可见部分腺上皮脱落,结构不清;其中以Gen高剂量组效果最明显,与对照组比较,Caspase-8在Gen治疗组中的表达率明显升高(P<0.05);Caspase-8在Gen高剂量组中的阳性表达率为80.0%,明显高于Gen低剂量组的69.2%,差异有统计学意义(P<0.01).与对照组比较,治疗组中NF-kB表达率无明显差异(P>0.05).结论:在经Gen处理后的大鼠EMs中,Gen诱导Caspase-8调节细胞凋亡,表明Caspase-8通过调节细胞凋亡参与了Gen治疗子宫内膜异位症的发生、发展过程.Gen对大鼠EMs具有良好的抑制其组织增生、促进细胞凋亡的作用,其作用机制可能是通过NF-kB以外的传导途径对EMs发挥作用.

  14. DNMT1 and DNMT3b silencing sensitizes human hepatoma cells to TRAIL-mediated apoptosis via up-regulation of TRAIL-R2/DR5 and caspase-8.

    Science.gov (United States)

    Kurita, Satoshi; Higuchi, Hajime; Saito, Yoshimasa; Nakamoto, Nobuhiro; Takaishi, Hiromasa; Tada, Shinichiro; Saito, Hidetsugu; Gores, Gregory J; Hibi, Toshifumi

    2010-06-01

    DNA methylation plays a critical role in chromatin remodeling and gene expression. DNA methyltransferases (DNMTs) are hypothesized to mediate cellular DNA methylation status and gene expression during mammalian development and in malignant diseases. In this study, we examined the role of DNA methyltransferase 1 (DNMT1) and DNMT3b in cell proliferation and survival of hepatocellular carcinoma (HCC) cells. Gene silencing of both DNMT1 and DNMT3b by targeted siRNA knockdown reduces cell proliferation and sensitizes the cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated cell death. The proapoptotic protein caspase-8 demonstrated promoter hypermethylation in HCC cells and was up-regulated by knockdown of DNMT1 and DNMT3b both at mRNA and protein levels. In addition, death receptor TRAIL-R2/DR5 (TRAIL receptor 2/death receptor 5) did not exhibit promoter hypermethylation in HCC cells but was also up-regulated by knockdown of DNMT1 and DNMT3b both at mRNA and protein levels. Consistent with this observation, the combined transfection of DNMT1-siRNA plus DNMT3b-siRNA enhanced formation of the TRAIL-death-inducing signaling complex formation in HCC cells. In conclusion, our data suggest that DNA methylation of specific genomic regions maintained by DNMT1 and DNMT3b plays a critical role in survival of HCC cells, and a simultaneous knockdown of both DNMT1 and DNMT3b may be a novel anticancer strategy for the treatment of HCC.

  15. 半胱氨酸蛋白酶-8和P53在慢性砷中毒大鼠肾近端小管表达的研究%Expression of cysteine caspase-8 and P53 in renal proximal tubular epithelial cell of chronic arsenic poisoning rats

    Institute of Scientific and Technical Information of China (English)

    钱立全; 李远慧; 孔祥照; 金婷婷; 李娜

    2012-01-01

    Objective To study the molecular mechanism of renal injury of chronic arsenic poisoning rats induced by the expression of cysteine caspase-8 and P53 in renal proximal tubular epithelial cells.Methods Sixty healthy SD rats were divided into three groups,high-,low-dose group,and control group,n =20 in each group.The rats in high and low dose groups were treated with As203 through drinking water,10.0 and 0.4 mg/kg,respectively.The control rats were given distilled water.Four months later,serum and urinary arsenic level was determined,and kidney specimens were taken.The expression of cysteine caspase-8 and P53 in renal proximal tubular epithelial cells was detected by histological technique-HE staining and SABC immunohistochemistry.In addition,cell number counting and image analyses were used in the study.Results The number of caspase-8 positive cells of renal proximal tubule in control group,low-and high-dose group was 3.33±1.32,31.14±8.02 and 46.50±7.20 cell number/visual fields,respectively,which was increased with dose increasing(all P <0.05);the average gray value was 151.34±6.40,133.58±4.63 and 128.34±16.28,respectively,decreased with dose increasing(all P <0.05).The number of P53 positive cells was 3.17±1.59,26.29±4.23 and 47.00±6.22 cell number/visual fields,respectively,increased with dose increasing (all P < 0.05) ; the average gray value was 142.54±8.06,121.48±5.68 and 101.89±6.35,respectively,decreased with dose increasing (all P < 0.05).Conclusion The increase of caspase-8 and P53 positive cells is one of the molecular mechanisms of renal injury induced by arsenic poisoning.%目的 探讨慢性砷中毒大鼠肾近端小管细胞半胱氨酸蛋白酶-8(caspase-8)和P53表达致肾脏损伤的分子机制.方法 60只SD大鼠分为对照组,低、高剂量组,每组20只,高、低剂量组分别给予三氧化二砷(As2O3) 10.0、0.4 mg/kg水溶液自由饮用,对照组自由饮用蒸馏水.分笼4个月,测定血砷、尿砷,取肾脏

  16. 携SH2-Caspase8融合基因重组腺病毒的构建及对K562细胞增殖的影响%Construction of recombinant adenovirus carrying SH2-Caspase8 fusion gene and its effect on proliferation in K562 cells

    Institute of Scientific and Technical Information of China (English)

    刘双凤; 胡晶; 曹唯希; 史静; 彭智; 王海霞; 李亚娟; 冯文莉

    2011-01-01

    Objective To construct the recombinant adenovirus carrying SH2-Caspase8 fusion gene and its mutant to observe its inhibitory effect on proliferation of K562 cells. Methods SH2-Caspase8 fusion gene was amplified by RT-PCR and splicing PCR, and cloned into the shuttle plasmid pAdTrack-CMV of adenovirus. The shuttle plasmid pAdT-SC-EGFP was constructed and identified by double-enzyme digestion and DNA sequencing, and transformed into ultra-competent pAdEasy-BJ5183 after Pme Ⅰ digestion. Defective replication recombinant adenovirus plasmid pAdE-SC-EGFP and its mutant were generated by homologous recombination and further transfected into AD293 cells to package recombinant adenovirus after Pac Ⅰ digestion. Recombinant adenovirus plasmid AdE-SC-EGFP was identified by PCR and Western blotting, respectively, and amplified for the measurement of its titers. Results PCR, enzyme digestion and DNA sequencing showed that the shuttle and recombinant plasmids of adenovirus, pAdT-SC-EGFP and pAdE-SC-EGFP, were successfully constructed. PCR and EGFP expression confirmed that the recombinant plasmid of adenovirus, pAdE-SC-EGFP, was successfully packaged. After amplification, its titer was 1.5 ×l012pfu/ml. Western blot analysis displayed the expression of target protein. MTT and colony formation test could inhibit the proliferation of K562 cells. Conclusion Recombinant adenovirus plasmid, AdE-SC-EGFP, carrying SH2-Caspase8 fusion gene and its mutant, has been successfully constructed, and can significantly inhibit the proliferation of BCR-ABL positive K562 cells in vitro.%目的构建并鉴定携SH2 -Caspase8融合基因的重组腺病毒AdE-SC-EGFP及其突变体,观察其对K562细胞增殖的抑制作用。方法采用RT-PCR、重叠PCR扩增SH2-Caspase8融合基因,克隆至腺病毒穿梭载体pAdTrack-CMV中,构建穿梭质粒pAdT-SC-EGFP,进行酶切和测序鉴定。将PmeⅠ酶切后的穿梭质粒转化pAdEasy-BJ5183感受态,通过细菌内同源重组产

  17. Calpains participate in nerve terminal degeneration induced by spider and snake presynaptic neurotoxins.

    Science.gov (United States)

    Duregotti, Elisa; Tedesco, Erik; Montecucco, Cesare; Rigoni, Michela

    2013-03-15

    α-latrotoxin and snake presynaptic phospholipases A2 neurotoxins target the presynaptic membrane of axon terminals of the neuromuscular junction causing paralysis. These neurotoxins display different biochemical activities, but similarly alter the presynaptic membrane permeability causing Ca(2+) overload within the nerve terminals, which in turn induces nerve degeneration. Using different methods, here we show that the calcium-activated proteases calpains are involved in the cytoskeletal rearrangements that we have previously documented in neurons exposed to α-latrotoxin or to snake presynaptic phospholipases A2 neurotoxins. These results indicate that calpains, activated by the massive calcium influx from the extracellular medium, target fundamental components of neuronal cytoskeleton such as spectrin and neurofilaments, whose cleavage is functional to the ensuing nerve terminal fragmentation.

  18. Recombinant N-Terminal Slit2 Inhibits TGF-β-Induced Fibroblast Activation and Renal Fibrosis.

    Science.gov (United States)

    Yuen, Darren A; Huang, Yi-Wei; Liu, Guang-Ying; Patel, Sajedabanu; Fang, Fei; Zhou, Joyce; Thai, Kerri; Sidiqi, Ahmad; Szeto, Stephen G; Chan, Lauren; Lu, Mingliang; He, Xiaolin; John, Rohan; Gilbert, Richard E; Scholey, James W; Robinson, Lisa A

    2016-09-01

    Fibrosis and inflammation are closely intertwined injury pathways present in nearly all forms of CKD for which few safe and effective therapies exist. Slit glycoproteins signaling through Roundabout (Robo) receptors have been described to have anti-inflammatory effects through regulation of leukocyte cytoskeletal organization. Notably, cytoskeletal reorganization is also required for fibroblast responses to TGF-β Here, we examined whether Slit2 also controls TGF-β-induced renal fibrosis. In cultured renal fibroblasts, which we found to express Slit2 and Robo-1, the bioactive N-terminal fragment of Slit2 inhibited TGF-β-induced collagen synthesis, actin cytoskeletal reorganization, and Smad2/3 transcriptional activity, but the inactive C-terminal fragment of Slit2 did not. In mouse models of postischemic renal fibrosis and obstructive uropathy, treatment with N-terminal Slit2 before or after injury inhibited the development of renal fibrosis and preserved renal function, whereas the C-terminal Slit2 had no effect. Our data suggest that administration of recombinant Slit2 may be a new treatment strategy to arrest chronic injury progression after ischemic and obstructive renal insults by not only attenuating inflammation but also, directly inhibiting renal fibrosis.

  19. Chlorella vulgaris triggers apoptosis in hepatocarcinogenesis-induced rats

    Institute of Scientific and Technical Information of China (English)

    Emey Suhana MOHD AZAMAI; Suhaniza SULAIMAN; Shafina Hanim MOHD HABIB; Mee Lee LOOI; Srijit DAS; Nor Aini ABDUL HAMID; Wan Zurinah WANG NGAH; Yasmin Anum MOHD YUSOF

    2009-01-01

    Chlorella vulgaris (CV) has been reported to have antioxidant and anticancer properties. We evaluated the effect of CV on apoptotic regulator protein expression in liver cancer-induced rats. Male Wistar rats (200-250 g) were divided into eight groups: control group (normal diet), CDE group (choline deficient diet supplemented with ethionine in drinking water to induce hepatocarcinogenesis), CV groups with three different doses of CV (50, 150, and 300 mg/kg body weight), and CDE groups treated with different doses of CV (50, 150, and 300 mg/kg body weight). Rats were sacrificed at various weeks and liver tissues were embedded in paraffin blocks for immunohistochemistry studies. CV, at increasing doses, decreased the expression of anti-apoptotic protein, Bcl-2, but increased the expression of pro-apoptotic protein, caspase 8, in CDE rats, which was correlated with decreased hepatoctyes proliferation and increased apoptosis as determined by bromodeoxy-uridine (BrdU) labeling and terminal deoxynucleotidyl transferase mediated dUTP nick-end labeling (TUNEL) assay, respectively. Our study shows that CV has definite chemopreventive effect by inducing apoptosis via decreasing the expression of Bcl-2 and increasing the expression of caspase 8 in hepatocarcinogenesis-induced rats.

  20. PI3-kinase-dependent activation of apoptotic machinery oc-curs on commitment of epidermal keratinocytes to terminal differentiation

    Institute of Scientific and Technical Information of China (English)

    Sam M Janes; Tyler A Ofstad; Douglas H Campbell; Ayad Eddaoudi; Gary Warnes; Derek Davies; Fiona M Watt

    2009-01-01

    We have investigated the earliest events in commitment of human epidermal keratinocytes to terminal differen-tiation. Phosphorylated Akt and caspase activation were detected in cells exiting the basal layer of the epidermis. Activation of Akt by retroviral transduction of primary cultures of human keratinocytes resulted in an increase in abortive clones founded by transit amplifying cells, while inhibition of the upstream kinase, Pl3-kinase, inhibited suspension-induced terminal differentiation. Caspase inhibition also blocked differentiation, the primary mediator being caspase 8. Caspase activation was initiated by 2 h in suspension, preceding the onset of expression of the termi-nal differentiation marker involucrin by several hours. Incubation of suspended cells with fibronectin or inhibition of PI3-kinase prevented caspase induction. At 2 h in suspension, keratinocytes that had become committed to terminal differentiation had increased side scatter, were 7-aminoactinomycin D (7-AAD) positive and annexin V negative; they exhibited loss of mitochondrial membrane potential and increased cardiolipin oxidation, but with no increase in reac-tive oxygen species. These properties indicate that the onset of terminal differentiation, while regulated by Pl3-kinase and caspases, is not a classical apoptotic process.

  1. Selective presynaptic terminal remodeling induced by spatial, but not cued, learning: a quantitative confocal study.

    Science.gov (United States)

    McGonigal, R; Tabatadze, N; Routtenberg, A

    2012-06-01

    The hippocampal mossy fibers (MFs) are capable of behaviorally selective, use-dependent structural remodeling. Indeed, we previously observed a new layer of Timm's staining induced in the stratum oriens (SO) in CA3 after spatial but not cued water maze learning (Rekart et al., (2007) Learn Mem 14:416-421). This led to the prediction that there is a learning-specific induction of presynaptic terminal plasticity of MF axons. This study confirms this prediction demonstrating, at the confocal level of analysis, terminal-specific, and behavior-selective presynaptic structural plasticity linked to long-term memory. Male adult Wistar rats were trained for 5 days to locate a hidden or visible platform in a water maze and a retention test was performed 7 days later. MF terminal subtypes, specifically identified by an antibody to zinc transporter 3 (ZnT3), were counted from confocal z-stacks in the stratum lucidum (SL) and the SO. In hidden platform trained rats, there was a significant increase in the number of large MF terminals (LMTs, 2.5-10 μm diameter, >2 μm(2) area) compared to controls both in the proximal SL (P CA3 pyramidal neurons, while SMTs are known to target inhibitory interneurons. The present findings highlight the pivotal role in memory of presynaptic structural plasticity. Because the "sprouting" observed is specific to the LMT, with no detectable change in the number of the SMT, learning may enhance net excitatory input to CA3 pyramidal neurons. Given the sparse coding of the MF-CA3 connection, and the role that granule cells play in pattern separation, the remodeling observed here may be expected to have a major impact on the long-term integration of spatial context into memory.

  2. Selective presynaptic terminal remodeling induced by spatial, but not cued, learning: a quantitative confocal study

    Science.gov (United States)

    McGonigal, R.; Tabatadze, N.; Routtenberg, A.

    2011-01-01

    The hippocampal mossy fibers (MFs) are capable of behaviorally-selective, use-dependent structural remodeling. Indeed, we previously observed a new layer of Timm’s staining induced in the stratum oriens (SO) in CA3 after spatial but not cued water maze learning (Rekart et al., Learn. Mem. 2007; 14:416–421). This led to the prediction that there is a learning-specific induction of presynaptic terminal plasticity of MF axons. The present study confirms this prediction demonstrating, at the confocal level of analysis, terminal-specific and behavior-selective presynaptic structural plasticity linked to long-term memory. Male adult Wistar rats were trained for 5d to locate a hidden or visible platform in a water maze and a retention test was performed 7d later. MF terminal subtypes, specifically identified by an antibody to zinc transporter 3 (ZnT3), were counted from confocal z-stacks in the stratum lucidum (SL) and the SO. In hidden platform trained rats there was a significant increase in the number of large MF terminals (LMTs, 2.5–10µm diameter, >2µm2 area) compared to controls both in the proximal SL (p CA3 pyramidal neurons, while SMTs are known to target inhibitory interneurons. The present findings highlight the pivotal role in memory of presynaptic structural plasticity. Because the ‘sprouting’ observed is specific to the LMT, with no detectable change in the number of the SMT, learning may enhance net excitatory input to CA3 pyramidal neurons. Given the sparse coding of the MF-CA3 connection, and the role that granule cells play in pattern separation, the remodeling observed here may be expected to have a major impact on the long-term integration of spatial context into memory. PMID:22180136

  3. Charge-exchange Induced Modulation of the Heliosheath Ion Distribution Downstream of the Termination Shock

    Science.gov (United States)

    Fahr, H. J.; Fichtner, H.; Scherer, K.

    2015-12-01

    We consider the evolution of the solar wind ion distribution function alongthe plasma flow downstream from the termination shock induced by chargeexchange processes with cold interstellar H-atoms. We start from a kineticphase space transport equation valid in the bulk frame of the plasma flowthat takes into account convective changes, cooling processes, energydiffusion and ion injection, and describes solar wind and pick-up ionsas a co-moving, isotropic, joint ion population. From this kinetic transportequation one can ascend to an equation for the pressure moment of the iondistribution function, a so-called pressure transport equation, describingthe evolution of the ion pressure in the comoving rest frame. Assuming thatthe local ion distribution can be represented by an adequate kappa functionwith a kappa parameter that varies with the streamline coordinate, weobtain an ordinary differential equation for kappa as function of thestreamline coordinate s. With this result then we gain the heliosheath iondistribution function downstream of the termination shock. The latter thencan be used to predict the Voyager-2 measured moments of the distributionfunction like ion density and ion temperature, and it can also be used topredict spectral fluxes of ENA`s originating from these ions and registeredby IBEX-Hi and IBEX-Lo.We especially analyse the solar wind ion temperature decreasemeasured by Voyager-2 between the years 2008 to 2011 and try to explain itas a charge-exchange induced cooling of the ion distribution function duringthe associated ion convection period.

  4. Cell autonomy of HIF effects in Drosophila: tracheal cells sense hypoxia and induce terminal branch sprouting.

    Science.gov (United States)

    Centanin, Lázaro; Dekanty, Andrés; Romero, Nuria; Irisarri, Maximiliano; Gorr, Thomas A; Wappner, Pablo

    2008-04-01

    Drosophila tracheal terminal branches are plastic and have the capacity to sprout out projections toward oxygen-starved areas, in a process analogous to mammalian angiogenesis. This response involves the upregulation of FGF/Branchless in hypoxic tissues, which binds its receptor Breathless on tracheal cells. Here, we show that extra sprouting depends on the Hypoxia-Inducible Factor (HIF)-alpha homolog Sima and on the HIF-prolyl hydroxylase Fatiga that operates as an oxygen sensor. In mild hypoxia, Sima accumulates in tracheal cells, where it induces breathless, and this induction is sufficient to provoke tracheal extra sprouting. In nontracheal cells, Sima contributes to branchless induction, whereas overexpression of Sima fails to attract terminal branch outgrowth, suggesting that HIF-independent components are also required for full induction of the ligand. We propose that the autonomous response to hypoxia that occurs in tracheal cells enhances tracheal sensitivity to increasing Branchless levels, and that this mechanism is a cardinal step in hypoxia-dependent tracheal sprouting.

  5. Downregulation of active caspase 8 as a mechanism of acquired TRAIL resistance in mismatch repair-proficient colon carcinoma cell lines

    NARCIS (Netherlands)

    Van Geelen, Caroline M. M.; Pennarun, Bodvael; Boerma-Van Ek, Wytske; Le, Phuong T. K.; Spierings, Diana C. J.; De Vries, Elisabeth G. E.; De Jong, Steven

    2010-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) triggers the apoptotic cascade in various colon cancer cell lines after binding to the membrane receptors DR4 and DR5. However, not all cancer cell lines are sensitive to the therapeutic recombinant human TRAIL (rhTRAIL). To investigate

  6. COMBINATION OF γ-INTERFERON WITH TRAIL AND CISPLATIN OR ETOPOSIDE INDUCES APOPTOSIS IN HUMAN NEUROBLASTOMA CELL LINE SH-SY5Y

    Institute of Scientific and Technical Information of China (English)

    Hai-xia Tong; Chun-wei Lu; Ji-hong Zhang; Li Ma; Jin-hua Zhang

    2007-01-01

    Objective To study the effect of γ-interferon (IFN-γ), tumor necrosis factor related apoptosis inducing ligand (TRAIL), and cisplatin or etoposide induced apoptosis in human neuroblastoma cell line SH-SY5Y and its possible molecular mechanisms.Methods The expressions of Caspase 8 mRNA and protein were detected with RT-PCR and Western blot analysis. The effects of IFNγ, TRAIL, IFNγ + TRAIL, IFNγ + Caspase 8 inhibitor + TRAIL, IFNγ + cisplatin + TRAIL, and IFNγ + etoposide + TRAIL on the growth and apoptosis of SH-SY5Y cells were detected with the methods of MTT and flow cytometry. The relative Caspase 8 activity was measured with colorimetric assay.Results Caspase 8 was undetectable in SH-SY5Y cells but an increased expression of Caspase 8 mRNA and protein was found after treatment with IFNγ. SH-SY5Y cells themselves were not sensitive to TRAIL, but those expressing Caspase 8 after treatment with IFNγ were. The killing effect of TRAIL on SH-SY5 Y cells expressing Caspase 8 was depressed by Caspase 8 inhibitor. Cisplatin and etoposide could enhance the sensitivity of TRAIL on SH-SY5Y cells. The relative Caspase 8 activity of SH-SY5Y cells in IFNγ + TRAIL group was significantly higher than those of control group, IFNγ group, TRAIL group, and inhibitor group (P <0.01). There was no significant difference among IFNγ + TRAIL group, IFNγ + cisplatin + TRAIL group, and IFNγ + etoposide + TRAIL group.Conclusions IFNγ could sensitize SH-SY5Y cells to TRAIL-induced apoptosis and this may be realized by the up-regulation of Caspase 8. Cisplatin and etoposide could enhance the killing effect of TRAIL on SH-SY5Y cells.

  7. Nested N-terminal megalin fragments induce high-titer autoantibody and attenuated Heymann nephritis.

    Science.gov (United States)

    Tramontano, Alfonso; Knight, Thomas; Vizzuso, Domenica; Makker, Sudesh P

    2006-07-01

    It was shown previously that an N-terminal fragment (nM60) that encompasses amino acid residues 1 to 563 of megalin could induce active Heymann nephritis (AHN) as efficiently as the native protein. For delineation of a minimal structure within this fragment that is sufficient to induce AHN, smaller protein fragments that encompass residues 1 to 236 (L6), 1 to 195 (L5), 1 to 156 (L4), and 1 to 120 (L3), representing successive C-terminal truncations within ligand-binding repeats of nM60, were cloned and produced in a baculovirus insect cell expression system. Protein fragments L4, L5, and L6 clearly were glycosylated. All four fragments stimulated proliferation of megalin-sensitized lymph node cells and induced high-titer anti-megalin autoantibodies in Lewis rats. A full-blown disease, as assessed by severity of proteinuria, was observed in rats that were immunized with L6 and L5, whereas animals that were immunized with L4 and L3 developed only mild disease. The proteinuria levels correlated with staining for complement (C3, C5b-9) and IgG1 isotype in glomerular immune deposits. The results suggest that one or more molecular determinants on the region that comprises amino acid residues 157 to 236 contribute to the induction of a full-blown form of AHN. Study of the structure, conformation, and posttranslational modifications of these determinants could provide greater insight into the molecular correlates of immunopathogenesis in this disease model.

  8. Screening for Small Molecule Inhibitors of Statin-Induced APP C-terminal Toxic Fragment Production

    Science.gov (United States)

    Poksay, Karen S.; Sheffler, Douglas J.; Spilman, Patricia; Campagna, Jesus; Jagodzinska, Barbara; Descamps, Olivier; Gorostiza, Olivia; Matalis, Alex; Mullenix, Michael; Bredesen, Dale E.; Cosford, Nicholas D. P.; John, Varghese

    2017-01-01

    Alzheimer’s disease (AD) is characterized by neuronal and synaptic loss. One process that could contribute to this loss is the intracellular caspase cleavage of the amyloid precursor protein (APP) resulting in release of the toxic C-terminal 31-amino acid peptide APP-C31 along with the production of APPΔC31, full-length APP minus the C-terminal 31 amino acids. We previously found that a mutation in APP that prevents this caspase cleavage ameliorated synaptic loss and cognitive impairment in a murine AD model. Thus, inhibition of this cleavage is a reasonable target for new therapeutic development. In order to identify small molecules that inhibit the generation of APP-C31, we first used an APPΔC31 cleavage site-specific antibody to develop an AlphaLISA to screen several chemical compound libraries for the level of N-terminal fragment production. This antibody was also used to develop an ELISA for validation studies. In both high throughput screening (HTS) and validation testing, the ability of compounds to inhibit simvastatin- (HTS) or cerivastatin- (validation studies) induced caspase cleavage at the APP-D720 cleavage site was determined in Chinese hamster ovary (CHO) cells stably transfected with wildtype (wt) human APP (CHO-7W). Several compounds, as well as control pan-caspase inhibitor Q-VD-OPh, inhibited APPΔC31 production (measured fragment) and rescued cell death in a dose-dependent manner. The effective compounds fell into several classes including SERCA inhibitors, inhibitors of Wnt signaling, and calcium channel antagonists. Further studies are underway to evaluate the efficacy of lead compounds – identified here using cells and tissues expressing wt human APP – in mouse models of AD expressing mutated human APP, as well as to identify additional compounds and determine the mechanisms by which they exert their effects.

  9. Screening for Small Molecule Inhibitors of Statin-Induced APP C-terminal Toxic Fragment Production.

    Science.gov (United States)

    Poksay, Karen S; Sheffler, Douglas J; Spilman, Patricia; Campagna, Jesus; Jagodzinska, Barbara; Descamps, Olivier; Gorostiza, Olivia; Matalis, Alex; Mullenix, Michael; Bredesen, Dale E; Cosford, Nicholas D P; John, Varghese

    2017-01-01

    Alzheimer's disease (AD) is characterized by neuronal and synaptic loss. One process that could contribute to this loss is the intracellular caspase cleavage of the amyloid precursor protein (APP) resulting in release of the toxic C-terminal 31-amino acid peptide APP-C31 along with the production of APPΔC31, full-length APP minus the C-terminal 31 amino acids. We previously found that a mutation in APP that prevents this caspase cleavage ameliorated synaptic loss and cognitive impairment in a murine AD model. Thus, inhibition of this cleavage is a reasonable target for new therapeutic development. In order to identify small molecules that inhibit the generation of APP-C31, we first used an APPΔC31 cleavage site-specific antibody to develop an AlphaLISA to screen several chemical compound libraries for the level of N-terminal fragment production. This antibody was also used to develop an ELISA for validation studies. In both high throughput screening (HTS) and validation testing, the ability of compounds to inhibit simvastatin- (HTS) or cerivastatin- (validation studies) induced caspase cleavage at the APP-D720 cleavage site was determined in Chinese hamster ovary (CHO) cells stably transfected with wildtype (wt) human APP (CHO-7W). Several compounds, as well as control pan-caspase inhibitor Q-VD-OPh, inhibited APPΔC31 production (measured fragment) and rescued cell death in a dose-dependent manner. The effective compounds fell into several classes including SERCA inhibitors, inhibitors of Wnt signaling, and calcium channel antagonists. Further studies are underway to evaluate the efficacy of lead compounds - identified here using cells and tissues expressing wt human APP - in mouse models of AD expressing mutated human APP, as well as to identify additional compounds and determine the mechanisms by which they exert their effects.

  10. KLF4 N-Terminal Variance Modulates Induced Reprogramming to Pluripotency

    Directory of Open Access Journals (Sweden)

    Shin-Il Kim

    2015-04-01

    Full Text Available As the quintessential reprogramming model, OCT3/4, SOX2, KLF4, and c-MYC re-wire somatic cells to achieve induced pluripotency. Yet, subtle differences in methodology confound comparative studies of reprogramming mechanisms. Employing transposons, we systematically assessed cellular and molecular hallmarks of mouse somatic cell reprogramming by various polycistronic cassettes. Reprogramming responses varied in the extent of initiation and stabilization of transgene-independent pluripotency. Notably, the cassettes employed one of two KLF4 variants, differing only by nine N-terminal amino acids, which generated dissimilar protein stoichiometry. Extending the shorter variant by nine N-terminal amino acids or augmenting stoichiometry by KLF4 supplementation rescued both protein levels and phenotypic disparities, implicating a threshold in determining reprogramming outcomes. Strikingly, global gene expression patterns elicited by published polycistronic cassettes diverged according to each KLF4 variant. Our data expose a Klf4 reference cDNA variation that alters polycistronic factor stoichiometry, predicts reprogramming hallmarks, and guides comparison of compatible public data sets.

  11. A Short Amino-Terminal Part of Arabidopsis Phytochrome A Induces Constitutive Photomorphogenic Response

    Institute of Scientific and Technical Information of China (English)

    András Viczián; (E)va (A)dám; Iris Wolf; János Bindics; Stefan Kircher; Marc Heijde; Roman UIm; Eberhard Sch(a)fer; Ferenc Nagy

    2012-01-01

    Phytochrome A (phyA) is the dominant photoreceptor of far-red light sensing in Arabidopsis thaliana.phyA accumulates at high levels in the cytoplasm of etiolated seedlings,and light-induced phyA signaling is mediated by a complex regulatory network.This includes light- and FHY1/FHL protein-dependent translocation of native phyA into the nucleus in vivo.It has also been shown that a short N-terminal fragment of phyA (PHYA406) is sufficient to phenocopy this highly regulated cellular process in vitro.To test the biological activity of this N-terminal fragment of phyA in planta,we produced transgenic phyA-201 plants expressing the PHYA406-YFP (YELLOW FLUORESCENT PROTEIN)-DD,PHYA406-YFP-DD-NLS (nuclear localization signal),and PHYA406-YFP-DD-NES (nuclear export signal) fusion proteins.Here,we report that PHYA406-YFP-DD is imported into the nucleus and this process is partially light-dependent whereas PHYA406-YFP-DD-NLS and PHYA406-YFP-DD-NES display the expected constitutive localization patterns.Our results show that these truncated phyA proteins are light-stable,they trigger a constitutive photomorphogenic-like response when localized in the nuclei,and neither of them induces proper phyA signaling.We demonstrate that in vitro and in vivo PHYA406 Pfr and Pr bind COP1,a general repressor of photomorphogenesis,and co-localize with it in nuclear bodies.Thus,we conclude that,in planta,the truncated PHYA406 proteins inactivate COP1 in the nuclei in a light-independent fashion.

  12. Interference of silibinin with IGF-1R signalling pathways protects human epidermoid carcinoma A431 cells from UVB-induced apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Weiwei; Otkur, Wuxiyar; Li, Lingzhi; Wang, Qiong; He, Hao; Zang, Linghe; Hayashi, Toshihiko [China–Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang 110016 (China); Tashiro, Shin-ichi [Institute for Clinical and Biomedical Sciences, Kyoto 603-8072 (Japan); Onodera, Satoshi [Department of Clinical and Biomedical Sciences, Showa Pharmaceutical University, Tokyo 194-8543 (Japan); Xia, Mingyu [China–Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang 110016 (China); Ikejima, Takashi, E-mail: ikejimat@vip.sina.com [China–Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang 110016 (China)

    2013-03-08

    Highlights: ► Silibinin protects A431 cells from UVB irradiation-induced apoptosis. ► Up-regulation of the IGF-1R-JNK/ERK pathways by UVB induces cell apoptosis. ► Silibinin inhibits IGF-1R pathways to repress caspase-8-mediated apoptosis. -- Abstract: Ultraviolet B (UVB) from sunlight is a major cause of cutaneous lesion. Silibinin, a traditional hepatic protectant, elicits protective effects against UVB-induced cellular damage. In A431 cells, the insulin-like growth factor-1 receptor (IGF-1R) was markedly up-regulated by UVB irradiation. The activation of the IGF-1R signalling pathways contributed to apoptosis of the cells rather than rescuing the cells from death. Up-regulated IGF-1R stimulated downstream mitogen-activated protein kinases (MAPKs), such as c-Jun N-terminal kinases (JNK) and extracellular signal-regulated protein kinases 1/2 (ERK1/2). The subsequent activation of caspase-8 and caspase-3 led to apoptosis. The activation of IGF-1R signalling pathways is the cause of A431 cell death. The pharmacological inhibitors and the small interfering RNA (siRNA) targeting IGF-1R suppressed the downstream activation of JNK/ERK-caspases to help the survival of the UVB-irradiated A431 cells. Indeed, silibinin treatment suppressed the IGF-1R-JNK/ERK pathways and thus protected the cells from UVB-induced apoptosis.

  13. Molecular Mechanism of Bovine Trabecular Meshwork Cells Apoptosis Induced by Dexamethasone and Protection by Pilocarpine

    Institute of Scientific and Technical Information of China (English)

    Yajuan Gu; Shujun Zeng; Pengxin Qiu; Yuping Wu; Dawei Peng; Guangmei Yan

    2005-01-01

    Purpose: To study the molecular mechanism of trabecular meshwork cells apoptosis induced by dexamethasone and the protection of pilocarpine.Methods: Determining mRNA expression with reverse transcription-polymerase chain reaction (RT-PCR), protein expression with Western blots and the percentage of apoptotic cells with fluorescent microscopy.Results: Dexamethasone up-regulated Fas proteins and affected Bax, caspase-8 and caspase-9 proteins in an action of first decrease then increase. Pre-treatment with pilocarpine decreased the four proteins expression, which were increased by dexamethasone. Pilocarpine self could decrease pro-apoptotic factors Bax, caspase-8 and caspase-9 proteins expression.Conclusion: Fas/FasL pathway participated in apoptotic process induced by dexamethasone in trabecular meshwork cells and the process was probably related with both caspase-8 and caspase-9 pathways. Pilocarpine protected the cells against apoptosis through down-regulating Fas, Bax, caspase-8 and caspase-9 proteins expression.

  14. Clinical, ultrasonography and haematology of aglepristone-induced mid-gestation pregnancy terminations in rabbits

    Directory of Open Access Journals (Sweden)

    Gözde R. Özalp

    2013-02-01

    Full Text Available Aglepristone is a safe abortifacient in cats, dogs and rabbits. Although no serious side effects have been reported, there is no information available about the effects of the medicine on haematological parameters. For the first time clinical and ultrasonographic features and haematological profiles were evaluated in rabbits treated with aglepristone 15 and 16 days after mating. Ten healthy 10–14 month-old New Zealand White female rabbits were mated with fertile bucks and pregnancies were confirmed by ultrasound 15 days later. Of these, 5 does were treated with aglepristone (test group, n = 5 whilst the remaining five (control group, n = 5 were treated with a saline solution (0.9% NaCl. The treatment dose was 10 mg⁄kg body weight, administered subcutaneously once daily on two consecutive days (day 15 and 16 post mating. Ultrasonographic, clinical and haematological assessments were performed daily. Aglepristone treatment induced embryonic fluid resorptions without foetal death in mid-gestation terminations. Following ultrasonographic and haematological examinations, it was established that aglepristone is a safe abortifacient in rabbits.

  15. Clinical, ultrasonography and haematology of aglepristone-induced mid-gestation pregnancy terminations in rabbits

    Directory of Open Access Journals (Sweden)

    Gözde R. Özalp

    2013-05-01

    Full Text Available Aglepristone is a safe abortifacient in cats, dogs and rabbits. Although no serious side effects have been reported, there is no information available about the effects of the medicine on haematological parameters. For the first time clinical and ultrasonographic features and haematological profiles were evaluated in rabbits treated with aglepristone 15 and 16 days after mating. Ten healthy 10–14 month-old New Zealand White female rabbits were mated with fertile bucks and pregnancies were confirmed by ultrasound 15 days later. Of these, 5 does were treated with aglepristone (test group, n = 5 whilst the remaining five (control group, n = 5 were treated with a saline solution (0.9% NaCl. The treatment dose was 10 mg⁄kg body weight, administered subcutaneously once daily on two consecutive days (day 15 and 16 post mating. Ultrasonographic, clinical and haematological assessments were performed daily. Aglepristone treatment induced embryonic fluid resorptions without foetal death in mid-gestation terminations. Following ultrasonographic and haematological examinations, it was established that aglepristone is a safe abortifacient in rabbits.

  16. Active protein aggregates induced by terminally attached self-assembling peptide ELK16 in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Zhou Bihong

    2011-02-01

    Full Text Available Abstract Background In recent years, it has been gradually realized that bacterial inclusion bodies (IBs could be biologically active. In particular, several proteins including green fluorescent protein, β-galactosidase, β-lactamase, alkaline phosphatase, D-amino acid oxidase, polyphosphate kinase 3, maltodextrin phosphorylase, and sialic acid aldolase have been successfully produced as active IBs when fused to an appropriate partner such as the foot-and-mouth disease virus capsid protein VP1, or the human β-amyloid peptide Aβ42(F19D. As active IBs may have many attractive advantages in enzyme production and industrial applications, it is of considerable interest to explore them further. Results In this paper, we report that an ionic self-assembling peptide ELK16 (LELELKLK2 was able to effectively induce the formation of cytoplasmic inclusion bodies in Escherichia coli (E. coli when attached to the carboxyl termini of four model proteins including lipase A, amadoriase II, β-xylosidase, and green fluorescent protein. These aggregates had a general appearance similar to the usually reported cytoplasmic inclusion bodies (IBs under transmission electron microscopy or fluorescence confocal microscopy. Except for lipase A-ELK16 fusion, the three other fusion protein aggregates retained comparable specific activities with the native counterparts. Conformational analyses by Fourier transform infrared spectroscopy revealed the existence of newly formed antiparallel beta-sheet structures in these ELK16 peptide-induced inclusion bodies, which is consistent with the reported assembly of the ELK16 peptide. Conclusions This has been the first report where a terminally attached self-assembling β peptide ELK16 can promote the formation of active inclusion bodies or active protein aggregates in E. coli. It has the potential to render E. coli and other recombinant hosts more efficient as microbial cell factories for protein production. Our observation might

  17. Apoptosis-Inducing Activity of Marine Sponge Haliclona sp. Extracts Collected from Kosrae in Nonsmall Cell Lung Cancer A549 Cells

    Directory of Open Access Journals (Sweden)

    Woori Bae

    2015-01-01

    Full Text Available Although various anticancer drugs have been developed for the treatment of nonsmall cell lung cancer, chemotherapeutic efficacy is still limited. Natural products such as phytochemicals have been screened as novel alternative materials, but alternative funds such as marine bioresources remain largely untapped. Of these resources, marine sponges have undergone the most scrutiny for their biological activities, including antiinflammatory, antiviral, and anticancer properties. However, the biological mechanisms of the activities of these marine sponges are still unclear. We investigated the anticancer activity of marine sponges collected from Kosrae in Micronesia and examined their mechanisms of action using nonsmall cell lung cancer A549 cells as a model system. Of 20 specimens, the Haliclona sp. (KO1304-328 showed both dose- and time-dependent cytotoxicity. Further, methanol extracts of Haliclona sp. significantly inhibited cell proliferation and cell viability. A549 cells treated with Haliclona sp. demonstrated induced expression of c-Jun N-terminal kinase (JNK, p53, p21, caspase-8, and caspase-3. The percentage of apoptotic cells significantly increased in A549 cultures treated with Haliclona sp. These results indicate that Haliclona sp. induces apoptosis via the JNK-p53 pathway and caspase-8, suggesting that this marine sponge is a good resource for the development of drugs for treatment of nonsmall cell lung cancer.

  18. Cleavage-induced termination in U2 snRNA gene expression

    Energy Technology Data Exchange (ETDEWEB)

    Nabavi, Sadeq [Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada N1G 2W1 (Canada); Nazar, Ross N., E-mail: rnnazar@uoguelph.ca [Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada N1G 2W1 (Canada)

    2010-03-12

    The maturation of many small nuclear RNAs is dependent on RNase III-like endonuclease mediated cleavage, which generates a loading site for the exosome complex that trims the precursor at its 3' end. Using a temperature sensitive Pac1 nuclease, here we show that the endonuclease cleavage is equally important in terminating the transcription of the U2 snRNA in Schizosaccharomyces pombe. Using a temperature sensitive Dhp1p 5' {yields} 3' exonuclease, we demonstrate that it also is an essential component of the termination pathway. Taken together the results support a 'reversed torpedoes' model for the termination and maturation of the U2 snRNA; the Pac1 endonuclease cleavage provides entry sites for the 3' and 5' exonuclease activities, leading to RNA maturation in one direction and transcript termination in the other.

  19. Global analysis of induced transcription factors and cofactors identifies Tfdp2 as an essential coregulator during terminal erythropoiesis.

    Science.gov (United States)

    Chen, Cynthia; Lodish, Harvey F

    2014-06-01

    Key transcriptional regulators of terminal erythropoiesis, such as GATA-binding factor 1 (GATA1) and T-cell acute lymphocytic leukemia protein 1 (TAL1), have been well characterized, but transcription factors and cofactors and their expression modulations have not yet been explored on a global scale. Here, we use global gene expression analysis to identify 28 transcription factors and 19 transcriptional cofactors induced during terminal erythroid differentiation whose promoters are enriched for binding by GATA1 and TAL1. Utilizing protein-protein interaction databases to identify cofactors for each transcription factor, we pinpoint several co-induced pairs, of which E2f2 and its cofactor transcription factor Dp-2 (Tfdp2) were the most highly induced. TFDP2 is a critical cofactor required for proper cell cycle control and gene expression. GATA1 and TAL1 are bound to the regulatory regions of Tfdp2 and upregulate its expression and knockdown of Tfdp2 results in significantly reduced rates of proliferation as well as reduced upregulation of many erythroid-important genes. Loss of Tfdp2 also globally inhibits the normal downregulation of many E2F2 target genes, including those that regulate the cell cycle, causing cells to accumulate in S phase and resulting in increased erythrocyte size. Our findings highlight the importance of TFDP2 in coupling the erythroid cell cycle with terminal differentiation and validate this study as a resource for future work on elucidating the role of diverse transcription factors and coregulators in erythropoiesis.

  20. Death, dynamics and disorder: Terminating reentry in excitable media by dynamically-induced inhomogeneities

    Indian Academy of Sciences (India)

    Johannes Breur; Sitabhra Sinha

    2005-04-01

    Formation of feedback loops of excitation waves (reentrant circuit) around non-conducting ventricular scar tissue is a common cause of cardiac arrhythmias, such as ventricular tachycardia, often leading to death. This is typically treated by rapid stimulation from an implantable device (ICD). However, the mechanisms of reentry termination success and, more importantly, failure, are poorly understood. To study such mechanisms, we simulated pacing termination of reentry in a model of cardiac tissue having significant restitution and dispersion properties. Our results show that rapid pacing dynamically generates conduction inhomogeneities in the reentrant circuit, leading to successful pacing termination of tachycardia. The study suggests that more effective pacing algorithms can be designed by taking into account the role of such dynamical inhomogeneities.

  1. Lacosamide diminishes dryness-induced hyperexcitability of corneal cold sensitive nerve terminals.

    Science.gov (United States)

    Kovács, Illés; Dienes, Lóránt; Perényi, Kristóf; Quirce, Susana; Luna, Carolina; Mizerska, Kamila; Acosta, M Carmen; Belmonte, Carlos; Gallar, Juana

    2016-09-15

    Lacosamide is an anti-epileptic drug that is also used for the treatment of painful diabetic neuropathy acting through voltage-gated sodium channels. The aim of this work was to evaluate the effects of acute application of lacosamide on the electrical activity of corneal cold nerve terminals in lacrimo-deficient guinea pigs. Four weeks after unilateral surgical removal of the main lachrimal gland in guinea pigs, corneas were excised and superfused in vitro at 34°C for extracellular electrophysiological recording of nerve terminal impulse activity of cold thermosensitive nerve terminals. The characteristics of the spontaneous and the stimulus-evoked (cooling ramps from 34°C to 15°C) activity before and in presence of lacosamide 100µM and lidocaine 100µM were compared. Cold nerve terminals (n=34) recorded from dry eye corneas showed significantly enhanced spontaneous activity (8.0±1.1 vs. 5.2±0.7imp/s; Placosamide and lidocaine decreased spontaneous activity and peak response to cooling ramps significantly (Placosamide (P>0.05) to the irrigation fluid. In summary, the application of lacosamide results in a significant decrease of the augmented spontaneous activity and responsiveness to cold of corneal sensory nerves from tear-deficient animals. Based on these promising results we speculate that lacosamide might be used to reduce the hyperexcitability of corneal cold receptors caused by prolonged ocular surface dryness due to hyposecretory or evaporative dry eye disease.

  2. HIV gp120 induces, NF-kappaB dependent, HIV replication that requires procaspase 8.

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    Gary D Bren

    Full Text Available BACKGROUND: HIV envelope glycoprotein gp120 causes cellular activation resulting in anergy, apoptosis, proinflammatory cytokine production, and through an unknown mechanism, enhanced HIV replication. METHODOLOGY/PRINCIPAL FINDINGS: We describe that the signals which promote apoptosis are also responsible for the enhanced HIV replication. Specifically, we demonstrate that the caspase 8 cleavage fragment Caspase8p43, activates p50/p65 Nuclear Factor kappaB (NF-kappaB, in a manner which is inhibited by dominant negative IkappaBalpha. This caspase 8 dependent NF-kappaB activation occurs following stimulation with gp120, TNF, or CD3/CD28 crosslinking, but these treatments do not activate NF-kappaB in cells deficient in caspase 8. The Casp8p43 cleavage fragment also transactivates the HIV LTR through NF-kappaB, and the absence of caspase 8 following HIV infection greatly inhibits HIV replication. CONCLUSION/SIGNIFICANCE: Gp120 induced caspase 8 dependent NF-kappaB activation is a novel pathway of HIV replication which increases understanding of the biology of T-cell death, as well as having implications for understanding treatment and prevention of HIV infection.

  3. PKC phosphorylates residues in the N-terminal of the DA transporter to regulate amphetamine-induced DA efflux.

    Science.gov (United States)

    Wang, Qiang; Bubula, Nancy; Brown, Jason; Wang, Yunliang; Kondev, Veronika; Vezina, Paul

    2016-05-27

    The DA transporter (DAT), a phosphoprotein, controls extracellular dopamine (DA) levels in the central nervous system through transport or reverse transport (efflux). Multiple lines of evidence support the claim that PKC significantly contributes to amphetamine-induced DA efflux. Other signaling pathways, involving CaMKII and ERK, have also been shown to regulate DAT mediated efflux. Here we assessed the contribution of putative PKC residues (S4, S7, S13) in the N-terminal of the DAT to amphetamine-induced DA efflux by transfecting DATs containing different serine to alanine (S-A) point mutations into DA pre-loaded HEK-293 cells and incubating these cells in amphetamine (2μM). The effects of a S-A mutation at the non-PKC residue S12 and a threonine to alanine (T-A) mutation at the ERK T53 residue were also assessed for comparison. WT-DATs were used as controls. In an initial experiment, we confirmed that inhibiting PKC with Go6976 (130nM) significantly reduced amphetamine-induced DA efflux. In subsequent experiments, cells transfected with the S4A, S12A, S13A, T53A and S4,7,13A mutants showed a reduction in amphetamine-induced DA efflux similar to that observed with Go6976. Interestingly, cells transfected with the S7A mutant, identified by some as a PKC-PKA residue, showed unperturbed WT-DAT levels of amphetamine-induced DA efflux. These results indicate that phosphorylation by PKC of select residues in the DAT N-terminal can regulate amphetamine-induced efflux. PKC can act either independently or in concert with other kinases such as ERK to produce this effect.

  4. Phospholipase C-gamma1 is required for subculture-induced terminal differentiation of normal human oral keratinocytes.

    Science.gov (United States)

    Oh, Ju-Eun; Kook, Joong-Ki; Park, Kyung-Hee; Lee, Gene; Seo, Byoung-Moo; Min, Byung-Moo

    2003-04-01

    Serial subculture of primary normal human oral keratinocytes (NHOKs) to the post-mitotic stage induces terminal differentiation, which is in part linked to elevated levels of phospholipase C (PLC)-gamma1. Therefore, PLC-gamma1 may be involved in the signal transduction system that leads to the calcium regulation of subculture-induced keratinocyte differentiation. To test this hypothesis, the expression of PLC-gamma1 in primary NHOKs was blocked by transfecting cells with the antisense PLC-gamma1 cDNA construct. These cells demonstrated dramatic reductions in PLC-gamma1 protein and in the differentiation markers involucrin and transglutaminase following calcium exposure and an increase (15-20%) in in vitro life span versus empty vector-transfected cells. In addition, we established the ability of antisense PLC-gamma1 to block the serial subculture-induced rise in intracellular calcium. Similar observations were made following treatment with the specific PLC inhibitor U73122. These results indicate that the terminal differentiation of NHOKs by serial subculture is associated with PLC-gamma1, which mediates calcium regulation by mobilizing intracellular calcium.

  5. Thrombotic microangiopathies and acute kidney injury induced by artificial termination of pregnancy.

    Science.gov (United States)

    Wu, H; Zou, H B; Xu, Y; Zhang, L

    2014-01-01

    Thrombotic microangiopathy (TMA) is a rare, but potentially lethal condition requiring rapid recognition, diagnosis and initiation of therapy. Here, we present two cases of women with hemolytic anemia, thrombocytopenia and acute kidney injury shortly after surgical termination of pregnancy. Histological examination of their kidneys revealed endothelial cell swelling and luminal stenosis or fibrin-containing thrombi in the glomeruli and arterioles, which support the diagnosis of TMA. The patients were treated with hemodialysis, plasma infusion and corticosteroids with or without immunosuppressive agents. Three weeks after treatment, one patient was cured and symptoms of the other patient markedly improved. Reporting of more cases of TMA associated with surgical termination of pregnancy will provide further insights into this rare disease, possibly aiding in identifying risk factors and improving time to clinical diagnosis, treatment and prognosis.

  6. Shape-induced terminal differentiation of human epidermal stem cells requires p38 and is regulated by histone acetylation.

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    John T Connelly

    Full Text Available Engineered model substrates are powerful tools for examining interactions between stem cells and their microenvironment. Using this approach, we have previously shown that restricted cell adhesion promotes terminal differentiation of human epidermal stem cells via activation of serum response factor (SRF and transcription of AP-1 genes. Here we investigate the roles of p38 MAPK and histone acetylation. Inhibition of p38 activity impaired SRF transcriptional activity and shape-induced terminal differentiation of human keratinocytes. In addition, inhibiting p38 reduced histone H3 acetylation at the promoters of SRF target genes, FOS and JUNB. Although histone acetylation correlated with SRF transcriptional activity and target gene expression, treatment with the histone de-acetylase inhibitor, trichostatin A (TSA blocked terminal differentiation on micro-patterned substrates and in suspension. TSA treatment simultaneously maintained expression of LRIG1, TP63, and ITGB1. Therefore, global histone de-acetylation represses stem cell maintenance genes independent of SRF. Our studies establish a novel role for extrinsic physical cues in the regulation of chromatin remodeling, transcription, and differentiation of human epidermal stem cells.

  7. Heat shock-induced accumulation of translation elongation and termination factors precedes assembly of stress granules in S. cerevisiae.

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    Tomas Grousl

    Full Text Available In response to severe environmental stresses eukaryotic cells shut down translation and accumulate components of the translational machinery in stress granules (SGs. Since they contain mainly mRNA, translation initiation factors and 40S ribosomal subunits, they have been referred to as dominant accumulations of stalled translation preinitiation complexes. Here we present evidence that the robust heat shock-induced SGs of S. cerevisiae also contain translation elongation factors eEF3 (Yef3p and eEF1Bγ2 (Tef4p as well as translation termination factors eRF1 (Sup45p and eRF3 (Sup35p. Despite the presence of the yeast prion protein Sup35 in heat shock-induced SGs, we found out that its prion-like domain is not involved in the SGs assembly. Factors eEF3, eEF1Bγ2 and eRF1 were accumulated and co-localized with Dcp2 foci even upon a milder heat shock at 42°C independently of P-bodies scaffolding proteins. We also show that eEF3 accumulations at 42°C determine sites of the genuine SGs assembly at 46°C. We suggest that identification of translation elongation and termination factors in SGs might help to understand the mechanism of the eIF2α factor phosphorylation-independent repression of translation and SGs assembly.

  8. C-jun N-terminal Kinase-mediated Signaling Is Essential for Staphylococcus Aureus-induced U937 Apoptosis

    Institute of Scientific and Technical Information of China (English)

    Jia-he Wang; Bo Yu; Hui-yan Niu; Hui Li; Yi Zhang; Xin Wang; Ping He

    2009-01-01

    Objective To investigate the effect of SP600125, a specific c-jun N-terminal protein kinase (JNK) inhibitor, on Staphylococcus aureus (S. aureus)-induced U937 cell death and the underlying mechanism. Methods The human monocytic U937 cells were treated with S. aureus at different time with or without SP600125. Cell apoptosis was analyzed by flow cytometry. JNK, Bax, and caspase-3 activities were detected by Western blotting. Results S. aureus induced apoptosis in cultured U937 cells in a time-dependent manner. Expression of Bax and phospho-JNK significantly increased in S. aureus-treated U937 cells, and the level of activated caspase-3 also increased in a time-dependent manner. Inhibition of JNK with SP600125 significantly inhibited S. aureus-induced apoptosis in U937 cells. Conclusions S. aureus can induce apoptosis in U937 cells by phosphorylation of JNK and activation of Bax and caspase-3. SP600125 protects U937 cells from apoptosis induced by S. aureus via inhibiting the activity of JNK.

  9. Dicaffeoylquinic acids in Yerba mate (Ilex paraguariensis St. Hilaire) inhibit NF-kB nucleus translocation in macrophages and induce apoptosis by activating caspases-8 and -3 in human colon cancer cells

    Science.gov (United States)

    Caffeoylquinic acid derivatives (CQAs) are widely distributed in plants including Yerba mate (Ilex paraguariensis). However, isolation of these isomers in Yerba mate and their anti-inflammatory and anti-colon cancer effects has not been studied. The objectives of this study were to isolate and purif...

  10. Feedback activation of neurofibromin terminates growth factor-induced Ras activation

    OpenAIRE

    Hennig, Anne; Markwart, Robby; Wolff, Katharina; Schubert, Katja; Cui, Yan; Ian A Prior; Manuel A Esparza-Franco; Ladds, Graham; Rubio, Ignacio

    2016-01-01

    This is the final published version. It first appeared at http://biosignaling.biomedcentral.com/articles/10.1186/s12964-016-0128-z. Background Growth factors induce a characteristically short-lived Ras activation in cells emerging from quiescence. Extensive work has shown that transient as opposed to sustained Ras activation is critical for the induction of mitogenic programs. Mitogen-induced accumulation of active Ras-GTP results from increased nucleotide exchange driven by the nucleo...

  11. High-Fat-Diet-Induced Deficits in Dopamine Terminal Function Are Reversed by Restoring Insulin Signaling.

    Science.gov (United States)

    Fordahl, Steve C; Jones, Sara R

    2017-02-15

    Systemically released insulin crosses the blood-brain barrier and binds to insulin receptors on several neural cell types, including dopaminergic neurons. Insulin has been shown to decrease dopamine neuron firing in the ventral tegmental area (VTA), but potentiate release and reuptake at dopamine terminals in the nucleus accumbens (NAc). Here we show that prolonged consumption of a high fat diet blocks insulin's effects in the NAc, but insulin's effects are restored by inhibiting protein tyrosine phosphatase 1B, which supports insulin receptor signaling. Mice fed a high fat diet (60% kcals from fat) displayed significantly higher fasting blood glucose 160 mg/dL, compared to 101 mg/dL for control-diet-fed mice, and high-fat-diet-fed mice showed reduced blood glucose clearance after an intraperitoneal glucose tolerance test. Using fast scan cyclic voltammetry to measure electrically evoked dopamine in brain slices containing the NAc core, high-fat-diet-fed mice exhibited slower dopamine reuptake compared to control-diet-fed mice (2.2 ± 0.1 and 2.67 ± 0.15 μM/s, respectively). Moreover, glucose clearance rate was negatively correlated with Vmax. Insulin (10 nM to 1 μM) dose dependently increased reuptake rates in control-diet-fed mice compared with in the high-fat-diet group; however, the small molecule insulin receptor sensitizing agent, TCS 401 (300 nM), restored reuptake in high-fat-diet-fed mice to control-diet levels, and a small molecule inhibitor of the insulin receptor, BMS 536924 (300 nM), attenuated reuptake, similar to high-fat-diet-fed mice. These data show that a high-fat diet impairs dopamine reuptake by attenuating insulin signaling at dopamine terminals.

  12. Necroptosis suppresses inflammation via termination of TNF- or LPS-induced cytokine and chemokine production.

    Science.gov (United States)

    Kearney, C J; Cullen, S P; Tynan, G A; Henry, C M; Clancy, D; Lavelle, E C; Martin, S J

    2015-08-01

    TNF promotes a regulated form of necrosis, called necroptosis, upon inhibition of caspase activity in cells expressing RIPK3. Because necrosis is generally more pro-inflammatory than apoptosis, it is widely presumed that TNF-induced necroptosis may be detrimental in vivo due to excessive inflammation. However, because TNF is intrinsically highly pro-inflammatory, due to its ability to trigger the production of multiple cytokines and chemokines, rapid cell death via necroptosis may blunt rather than enhance TNF-induced inflammation. Here we show that TNF-induced necroptosis potently suppressed the production of multiple TNF-induced pro-inflammatory factors due to RIPK3-dependent cell death. Similarly, necroptosis also suppressed LPS-induced pro-inflammatory cytokine production. Consistent with these observations, supernatants from TNF-stimulated cells were more pro-inflammatory than those from TNF-induced necroptotic cells in vivo. Thus necroptosis attenuates TNF- and LPS-driven inflammation, which may benefit intracellular pathogens that evoke this mode of cell death by suppressing host immune responses.

  13. NFAT1 C-terminal domains are necessary but not sufficient for inducing cell death.

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    Douglas V Faget

    Full Text Available The proteins belonging to the nuclear factor of activated T cells (NFAT family of transcription factors are expressed in several cell types and regulate genes involved in differentiation, cell cycle and apoptosis. NFAT proteins share two conserved domains, the NFAT-homology region (NHR and a DNA-binding domain (DBD. The N- and C-termini display two transactivation domains (TAD-N and TAD-C that have low sequence similarity. Due to the high sequence conservation in the NHR and DBD, NFAT members have some overlapping roles in gene regulation. However, several studies have shown distinct roles for NFAT proteins in the regulation of cell death. The TAD-C shows low sequence similarity among NFAT family members, but its contribution to specific NFAT1-induced phenotypes is poorly understood. Here, we described at least two regions of NFAT1 TAD-C that confer pro-apoptotic activity to NFAT1. These regions extend from amino acids 699 to 734 and 819 to 850 of NFAT1. We also showed that the NFAT1 TAD-C is unable to induce apoptosis by itself and requires a functional DBD. Furthermore, we showed that when fused to NFAT1 TAD-C, NFAT2, which is associated with cell transformation, induces apoptosis in fibroblasts. Together, these results suggest that the NFAT1 TAD-C includes NFAT death domains that confer to different NFAT members the ability to induce apoptosis.

  14. [Pregnancy termination in Bulgaria – past, present and future perspectives. Drugs induced abortion – guidelines by WHO].

    Science.gov (United States)

    Marinov, B; Andreeva, A

    2013-01-01

    There are still too many unsafe abortions performed worldwide. Together with the efforts to reduce the abortion by choice, we note a rise in the need for mid trimester pregnancy termination for medical reasons. The article looks at the past present and future perspective of the abortion as a procedure in Bulgaria. States the fact that medical abortion is officially not widely performed. We reckon that with the existing guidelines by WHO and with Mifepriston and Misoprostol recently registered in Bulgaria, it is time for the medical abortion to become part of the clinical practice in Bulgaria. We believe that early medical abortion as well as mid trimester induced abortion is and adequate if not better alternative to the existing in Bulgaria procedures.

  15. Ligand-induced Ordering of the C-terminal Tail Primes STING for Phosphorylation by TBK1

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    Yuko Tsuchiya

    2016-07-01

    Full Text Available The innate immune protein Stimulator of interferon genes (STING promotes the induction of interferon beta (IFN-β production via the phosphorylation of its C-terminal tail (CTT by TANK-binding kinase 1 (TBK1. Potent ligands of STING are, therefore, promising candidates for novel anti-cancer drugs or vaccine adjuvants. However, the intrinsically flexible CTT poses serious problems in in silico drug discovery. Here, we performed molecular dynamics simulations of the STING fragment containing the CTT in ligand-bound and unbound forms and observed that the binding of a potent ligand cyclic GMP-AMP (cGAMP induced a local structure in the CTT, reminiscent of the known structure of a TBK1 substrate. The subsequent molecular biological experiments confirmed the observed dynamics of the CTT and identified essential residues for the activation of the IFN-β promoter, leading us to propose a new mechanism of STING activation.

  16. Mechanical effect of mélange-induced buttressing on embayment-terminating glacier dynamics

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    D. Seneca Lindsey

    2012-09-01

    Full Text Available Embayment terminating glaciers interact dynamically with seasonal sea ice and icebergs, a mixture we refer to as mélange. For certain glaciers, mélange prevents calved bergs from rotating away from the front, thus allowing the ice front to advance into the embayment. Here we demonstrate that mélange can, if rigid enough, provide sufficient buttressing to reduce the calving rate, while leaving the ice-front velocity largely unaffected. The net result is additional ice-front advance.

    Observations indicate a seasonal advance/retreat cycle has occurred at Jakobshavn Isbræ since the 1950s. We model an idealized Jakobshavn Isbræ-like scenario and find that mélange may be responsible for a seasonal ice-front advance of up to 0.6 km. These results come from a model that incorporates mélange into the interior of the domain, includes relevant stresses, and models drag via a kinematic boundary condition. A weakening or loss of mélange due to increasing temperatures would lead to further mass loss from glaciers such as Jakobshavn Isbræ.

  17. C-terminal of human histamine H1 receptors regulates their agonist-induced clathrin-mediated internalization and G-protein signaling.

    Science.gov (United States)

    Hishinuma, Shigeru; Nozawa, Hiroki; Akatsu, Chizuru; Shoji, Masaru

    2016-11-01

    It has been suggested that the agonist-induced internalization of G-protein-coupled receptors from the cell surface into intracellular compartments regulates cellular responsiveness. We previously reported that Gq/11 -protein-coupled human histamine H1 receptors internalized via clathrin-dependent mechanisms upon stimulation with histamine. However, the molecular determinants of H1 receptors responsible for agonist-induced internalization remain unclear. In this study, we evaluated the roles of the intracellular C-terminal of human histamine H1 receptors tagged with hemagglutinin (HA) at the N-terminal in histamine-induced internalization in Chinese hamster ovary cells. The histamine-induced internalization was evaluated by the receptor binding assay with [(3) H]mepyramine and confocal immunofluorescence microscopy with an anti-HA antibody. We found that histamine-induced internalization was inhibited under hypertonic conditions or by pitstop, a clathrin terminal domain inhibitor, but not by filipin or nystatin, disruptors of the caveolar structure and function. The histamine-induced internalization was also inhibited by truncation of a single amino acid, Ser487, located at the end of the intracellular C-terminal of H1 receptors, but not by its mutation to alanine. In contrast, the receptor-G-protein coupling, which was evaluated by histamine-induced accumulation of [(3) H]inositol phosphates, was potentiated by truncation of Ser487, but was lost by its mutation to alanine. These results suggest that the intracellular C-terminal of human H1 receptors, which only comprises 17 amino acids (Cys471-Ser487), plays crucial roles in both clathrin-dependent internalization of H1 receptors and G-protein signaling, in which truncation of Ser487 and its mutation to alanine are revealed to result in biased signaling toward activation of G-proteins and clathrin-mediated internalization, respectively.

  18. Identification of a Residue (Glu60) in TRAP Required for Inducing Efficient Transcription Termination at the trp Attenuator Independent of Binding Tryptophan and RNA.

    Science.gov (United States)

    McAdams, Natalie M; Patterson, Andrea; Gollnick, Paul

    2017-03-15

    Transcription of the tryptophan (trp) operon in Bacillus subtilis is regulated by an attenuation mechanism. Attenuation is controlled by the trpRNA-binding attenuation protein (TRAP). TRAP binds to a site in the 5' leader region of the nascent trp transcript in response to the presence of excess intracellular tryptophan. This binding induces transcription termination upstream of the structural genes of the operon. In prior attenuation models, the role of TRAP was only to alter the secondary structure of the leader region RNA so as to promote formation of the trp attenuator, which was presumed to function as an intrinsic terminator. However, formation of the attenuator alone has been shown to be insufficient to induce efficient termination, indicating that TRAP plays an additional role in this process. To further examine the function of TRAP, we performed a genetic selection for mutant TRAPs that bind tryptophan and RNA but show diminished termination at the trp attenuator. Five such TRAP mutants were obtained. Four of these have substitutions at Glu60, three of which are Lys (E60K) substitutions and the fourth of which is a Val (E60V) substitution. The fifth mutant obtained contains a substitution at Ile63, which is on the same β-strand of TRAP as Glu60. Purified E60K TRAP binds tryptophan and RNA with properties similar to those of the wild type but is defective at inducing termination at the trp attenuator in vitroIMPORTANCE Prior models for attenuation control of the B. subtilis trp operon suggested that the only role for TRAP is to bind to the leader region RNA and alter its folding to induce formation of an intrinsic terminator. However, several recent studies suggested that TRAP plays an additional role in the termination mechanism. We hypothesized that this function could involve residues in TRAP other than those required to bind tryptophan and RNA. Here we obtained TRAP mutants with alterations at Glu60 that are deficient at inducing termination in the

  19. Nematode-induced interference with vaccination efficacy targets follicular T helper cell induction and is preserved after termination of infection.

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    Irma Haben

    2014-09-01

    Full Text Available One-third of the human population is infected with parasitic worms. To avoid being eliminated, these parasites actively dampen the immune response of their hosts. This immune modulation also suppresses immune responses to third-party antigens such as vaccines. Here, we used Litomosoides sigmodontis-infected BALB/c mice to analyse nematode-induced interference with vaccination. Chronic nematode infection led to complete suppression of the humoral response to thymus-dependent vaccination. Thereby the numbers of antigen-specific B cells as well as the serum immunoglobulin (Ig G titres were reduced. TH2-associated IgG1 and TH1-associated IgG2 responses were both suppressed. Thus, nematode infection did not bias responses towards a TH2 response, but interfered with Ig responses in general. We provide evidence that this suppression indirectly targeted B cells via accessory T cells as number and frequency of vaccine-induced follicular B helper T cells were reduced. Moreover, vaccination using model antigens that stimulate Ig response independently of T helper cells was functional in nematode-infected mice. Using depletion experiments, we show that CD4+Foxp3+ regulatory T cells did not mediate the suppression of Ig response during chronic nematode infection. Suppression was induced by fourth stage larvae, immature adults and mature adults, and increased with the duration of the infection. By contrast, isolated microfilariae increased IgG2a responses to vaccination. This pro-inflammatory effect of microfilariae was overruled by the simultaneous presence of adults. Strikingly, a reduced humoral response was still observed if vaccination was performed more than 16 weeks after termination of L. sigmodontis infection. In summary, our results suggest that vaccination may not only fail in helminth-infected individuals, but also in individuals with a history of previous helminth infections.

  20. Nematode-Induced Interference with Vaccination Efficacy Targets Follicular T Helper Cell Induction and Is Preserved after Termination of Infection

    Science.gov (United States)

    Haben, Irma; Hartmann, Wiebke; Breloer, Minka

    2014-01-01

    One-third of the human population is infected with parasitic worms. To avoid being eliminated, these parasites actively dampen the immune response of their hosts. This immune modulation also suppresses immune responses to third-party antigens such as vaccines. Here, we used Litomosoides sigmodontis-infected BALB/c mice to analyse nematode-induced interference with vaccination. Chronic nematode infection led to complete suppression of the humoral response to thymus-dependent vaccination. Thereby the numbers of antigen-specific B cells as well as the serum immunoglobulin (Ig) G titres were reduced. TH2-associated IgG1 and TH1-associated IgG2 responses were both suppressed. Thus, nematode infection did not bias responses towards a TH2 response, but interfered with Ig responses in general. We provide evidence that this suppression indirectly targeted B cells via accessory T cells as number and frequency of vaccine-induced follicular B helper T cells were reduced. Moreover, vaccination using model antigens that stimulate Ig response independently of T helper cells was functional in nematode-infected mice. Using depletion experiments, we show that CD4+Foxp3+ regulatory T cells did not mediate the suppression of Ig response during chronic nematode infection. Suppression was induced by fourth stage larvae, immature adults and mature adults, and increased with the duration of the infection. By contrast, isolated microfilariae increased IgG2a responses to vaccination. This pro-inflammatory effect of microfilariae was overruled by the simultaneous presence of adults. Strikingly, a reduced humoral response was still observed if vaccination was performed more than 16 weeks after termination of L. sigmodontis infection. In summary, our results suggest that vaccination may not only fail in helminth-infected individuals, but also in individuals with a history of previous helminth infections. PMID:25255463

  1. Termination of Protofilament Elongation by Eribulin Induces Lattice Defects that Promote Microtubule Catastrophes.

    Science.gov (United States)

    Doodhi, Harinath; Prota, Andrea E; Rodríguez-García, Ruddi; Xiao, Hui; Custar, Daniel W; Bargsten, Katja; Katrukha, Eugene A; Hilbert, Manuel; Hua, Shasha; Jiang, Kai; Grigoriev, Ilya; Yang, Chia-Ping H; Cox, David; Horwitz, Susan Band; Kapitein, Lukas C; Akhmanova, Anna; Steinmetz, Michel O

    2016-07-11

    Microtubules are dynamic polymers built of tubulin dimers that attach in a head-to-tail fashion to form protofilaments, which further associate laterally to form a tube. Asynchronous elongation of individual protofilaments can potentially lead to an altered microtubule-end structure that promotes sudden depolymerization, termed catastrophe [1-4]. However, how the dynamics of individual protofilaments relates to overall growth persistence has remained unclear. Here, we used the microtubule targeting anti-cancer drug Eribulin [5-7] to explore the consequences of stalled protofilament elongation on microtubule growth. Using X-ray crystallography, we first revealed that Eribulin binds to a site on β-tubulin that is required for protofilament plus-end elongation. Based on the structural information, we engineered a fluorescent Eribulin molecule. We demonstrate that single Eribulin molecules specifically interact with microtubule plus ends and are sufficient to either trigger a catastrophe or induce slow and erratic microtubule growth in the presence of EB3. Interestingly, we found that Eribulin increases the frequency of EB3 comet "splitting," transient events where a slow and erratically progressing comet is followed by a faster comet. This observation possibly reflects the "healing" of a microtubule lattice. Because EB3 comet splitting was also observed in control microtubules in the absence of any drugs, we propose that Eribulin amplifies a natural pathway toward catastrophe by promoting the arrest of protofilament elongation.

  2. c-Jun N-terminal kinase is required for thermotherapy-induced apoptosis in human gastric cancer cells

    Institute of Scientific and Technical Information of China (English)

    Feng Xiao; Bin Liu; Qing-Xian Zhu

    2012-01-01

    AIM:To investigate the role of c-Jun N-terminal kinase (JNK) in thermotherapy-induced apoptosis in human gastric cancer SGC-7901 cells.METHODS:Human gastric cancer SGC-7901 cells were cultured in vitro.Following thermotherapy at 43 ℃ for 0,0.5,1,2 or 3 h,the cells were cultured for a further 24 h with or without the JNK specific inhibitor,SP600125 for 2 h.Apoptosis was evaluated by immunohistochemistry [terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)] and flow cytometry (Annexin vs propidium iodide).Cell proliferation was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide.The production of p-JNK,Bcl-2,Bax and caspase-3 proteins was evaluated by Western blotting.The expression of JNK at mRNA level was determined by reverse transcription polymerase chain reaction.RESULTS:The Proliferation of gastric carcinoma SGC-7901 cells was significantly inhibited following thermotherapy,and was 32.7%,30.6%,43.8% and 52.9% at 0.5,1,2 and 3 h post-thermotherapy,respectively.Flow cytometry analysis revealed an increased population of SGC-7901 cells in G0/G1 phase,but a reduced population in S phase following therrnotherapy for 1 or 2 h,compared to untreated cells (P < 0.05).The increased number of SGC-7901 cells in G0/G1 phase was consistent with induced apoptosis (flow cytometry) following thermotherapy for 0.5,1,2 or 3 h,compared to the untreated group (46.5% ± 0.23%,39.9% ± 0.53%,56.6% ±0.35% and 50.4% ± 0.29% vs 7.3% ± 0.10%,P < 0.01),respectively.This was supported by the TUNEL assay (48.2% ± 0.4%,40.1% ± 0.2%,61.2% ± 0.29% and 52.0% ± 0.42% vs 12.2% ± 0.22%,P < 0.01) respectively.More importantly,the expression of p-JNK protein and JNK mRNA levels were significantly higher at 0.5 h than at 0 h post-treatment (P < 0.01),and peaked at 2 h.A similar pattem was detected for Bax and caspase-3 proteins.Bcl-2 increased at 0.5 h,peaked at 1 h,and then decreased

  3. Inhibition of N-terminal lysines acetylation and transcription factor assembly by epirubicin induced deranged cell homeostasis.

    Directory of Open Access Journals (Sweden)

    Shahper N Khan

    Full Text Available Epirubicin (EPI, an anthracycline antitumour antibiotic, is a known intercalating and DNA damaging agent. Here, we study the molecular interaction of EPI with histones and other cellular targets. EPI binding with histone core protein was predicted with spectroscopic and computational techniques. The molecular distance r, between donor (histone H3 and acceptor (EPI was estimated using Förster's theory of non-radiation energy transfer and the detailed binding phenomenon is expounded. Interestingly, the concentration dependent reduction in the acetylated states of histone H3 K9/K14 was observed suggesting more repressed chromatin state on EPI treatment. Its binding site near N-terminal lysines is further characterized by thermodynamic determinants and molecular docking studies. Specific DNA binding and inhibition of transcription factor (Tf-DNA complex formation implicates EPI induced transcriptional inhibition. EPI also showed significant cell cycle arrest in drug treated cells. Chromatin fragmentation and loss of membrane integrity in EPI treated cells is suggestive of their commitment to cell death. This study provides an analysis of nucleosome dynamics during EPI treatment and provides a novel insight into its action.

  4. Drosophila DBT Autophosphorylation of Its C-Terminal Domain Antagonized by SPAG and Involved in UV-Induced Apoptosis.

    Science.gov (United States)

    Fan, Jin-Yuan; Means, John C; Bjes, Edward S; Price, Jeffrey L

    2015-07-01

    Drosophila DBT and vertebrate CKIε/δ phosphorylate the period protein (PER) to produce circadian rhythms. While the C termini of these orthologs are not conserved in amino acid sequence, they inhibit activity and become autophosphorylated in the fly and vertebrate kinases. Here, sites of C-terminal autophosphorylation were identified by mass spectrometry and analysis of DBT truncations. Mutation of 6 serines and threonines in the C terminus (DBT(C/ala)) prevented autophosphorylation-dependent DBT turnover and electrophoretic mobility shifts in S2 cells. Unlike the effect of autophosphorylation on CKIδ, DBT autophosphorylation in S2 cells did not reduce its in vitro activity. Moreover, overexpression of DBT(C/ala) did not affect circadian behavior differently from wild-type DBT (DBT(WT)), and neither exhibited daily electrophoretic mobility shifts, suggesting that DBT autophosphorylation is not required for clock function. While DBT(WT) protected S2 cells and larvae from UV-induced apoptosis and was phosphorylated and degraded by the proteasome, DBT(C/ala) did not protect and was not degraded. Finally, we show that the HSP-90 cochaperone spaghetti protein (SPAG) antagonizes DBT autophosphorylation in S2 cells. These results suggest that DBT autophosphorylation regulates cell death and suggest a potential mechanism by which the circadian clock might affect apoptosis.

  5. Roles of Fas signaling pathway in vitamin E succinate-induced apoptosis in human gastric cancer SGC-7901 cells

    Institute of Scientific and Technical Information of China (English)

    Kun Wu; Yao Li; Yan Zhao; Yu-Juan Shan; Wei Xia; Wei-Ping Yu; Lan Zhao

    2002-01-01

    AIM: To investigate the roles of Fas signaling pathway in vitamin E succinate-induced apoptosis in human gastric cancer SGC-7901 cells.METHODS: Human gastric cancer SGC-7901 cells were treated with VES at 5, 10, 20 mg@L-1, succinic acid and vitamin E as vehicle control and condition media only as untreated (UT) control. Apoptotic morphology was observed by DAPI staining. Western blot analysis was applied to measure the expression of Fas, FADD and caspase-8 proteins. After the cells were transiently transfected with Fas and FADD antisense oligonucleotides, respectively, caspase-8 activity was determined by flurometric method.RESULTS: The morphologically apoptotic changes were observed after VES treatment by DAPI staining. 23.7 % and 89.6 % apoptosis occurred after 24 h and 48 h of 20 mg@L-1 VES treatment, respectively. The protein levels of Fas, FADD and caspase-8 were evidently increased in a dose-dependent manner after 24 h of VES treatment. The blockage of Fas by transfection with Fas antisense oligonucleotides obviously inhibited the expression of FADD protein. After SGC-7901 cells were transfected with Fas and FADD antisense oligonucleotides, caspase-8 activity was obviously decreased (P<0.01), whereas Fas blocked more than FADD.CONCLUSION: VES-induced apoptosis in human gastric cancer SGC-7901 cells involves Fas signaling pathway including the interaction of Fas, FADD and caspase-8.

  6. Propolis from Turkey induces apoptosis through activating caspases in human breast carcinoma cell lines.

    Science.gov (United States)

    Seda Vatansever, H; Sorkun, Kadriye; Ismet Deliloğlu Gurhan, S; Ozdal-Kurt, Feyzan; Turkoz, Elgin; Gencay, Omur; Salih, Bekir

    2010-11-01

    Propolis is a sticky substance that is collected from plants by honeybees that has anti-mutagenic and anti-carcinogenic properties with biological and therapeutic effects. The target of this study was to investigate the anti-apoptotic effect of propolis extracts (PE) on the caspase pathway in the human breast cell line MCF-7 in culture. Seven different propolis extracts, numbered PE 1-7, produced in their natural ecological environment, were collected from the Hacettepe University Beytepe Campus area in Ankara, Turkey. Individual extracts at 0.5, 0.25, 0.125 and 0.063mg/ml were incubated with MCF-7 cells during 2 days culture. Cell growth and cytotoxicity were measured colorimetrically by MTT assay. Apoptotic cell death was determined by the TUNEL method (terminal deoxynucleotidyltransferase-biotin nick end-labelling) and caspase activity was investigated by immunocytochemistry using antibodies directed against caspase 6, caspase 8 and caspase 9. The results showed that the PE 5 and 6 extracts at 0.125mg/ml dilution induced apoptosis in association with increased number of TUNEL positive cells. MTT results showed that cultures exposed to the same extracts and at the same dilution experienced better cell growth compared to those cultures exposed to the other extracts. Immunpositivity for all caspases was detected after treatment with all the extracts and at all dilutions, with stronger immunoreactivity for caspase 6 than caspases 8 and 9. Caspase 6 labelling was especially strong in PE 5 and PE 6. We conclude that propolis may have anti-tumour effects by increasing apoptosis through the caspase pathway. Such propolis extracts may be important economically and allow development of a relatively inexpensive cancer treatment.

  7. Termination Documentation

    Science.gov (United States)

    Duncan, Mike; Hill, Jillian

    2014-01-01

    In this study, we examined 11 workplaces to determine how they handle termination documentation, an empirically unexplored area in technical communication and rhetoric. We found that the use of termination documentation is context dependent while following a basic pattern of infraction, investigation, intervention, and termination. Furthermore,…

  8. Left-handed helical preference in an achiral peptide chain is induced by an L-amino acid in an N-terminal type II β-turn.

    Science.gov (United States)

    De Poli, Matteo; De Zotti, Marta; Raftery, James; Aguilar, Juan A; Morris, Gareth A; Clayden, Jonathan

    2013-03-15

    Oligomers of the achiral amino acid Aib adopt helical conformations in which the screw-sense may be controlled by a single N-terminal residue. Using crystallographic and NMR techniques, we show that the left- or right-handed sense of helical induction arises from the nature of the β-turn at the N terminus: the tertiary amino acid L-Val induces a left-handed type II β-turn in both the solid state and in solution, while the corresponding quaternary amino acid L-α-methylvaline induces a right-handed type III β-turn.

  9. Protective Effect of Lupeol Against Lipopolysaccharide-Induced Neuroinflammation via the p38/c-Jun N-Terminal Kinase Pathway in the Adult Mouse Brain.

    Science.gov (United States)

    Badshah, Haroon; Ali, Tahir; Shafiq-ur Rehman; Faiz-ul Amin; Ullah, Faheem; Kim, Tae Hyun; Kim, Myeong Ok

    2016-03-01

    Recent studies have demonstrated a close interaction between neuroinflammatory responses, increased production of inflammatory mediators, and neurodegeneration. Pathological findings in neurological diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease have shown common signs of neuroinflammation and neurodegeneration. Lupeol, a natural pentacyclic triterpene, has revealed a number of pharmacological properties including an anti-inflammatory activity. This study aimed to evaluate the effect of lupeol against lipopolysaccharide (LPS)-induced neuroinflammation in the cortex and hippocampus of adult mice. Our results showed that systemic administration of LPS induced glial cell production of proinflammatory cytokines, tumor necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS), and interleukin (IL)-1β, while co-treatment with lupeol significantly inhibited the LPS-induced activation of microglia and astrocytes, and decreased the LPS-induced generation of TNF-α, iNOS, and IL-1β. The intracellular mechanism involved in the LPS-induced activation of inflammatory responses includes phosphorylation of P38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK), which was significantly inhibited by lupeol. We further elucidated that lupeol inhibited the LPS-induced activation of the mitochondrial apoptotic pathway and reversed the LPS-induced expression of apoptotic markers such as Bax, cytochrome C, caspase-9, and caspase-3. Taken together; our results suggest that lupeol inhibits LPS-induced microglial neuroinflammation via the P38-MAPK and JNK pathways and has therapeutic potential to treat various neuroinflammatory disorders.

  10. Analytically determined topological phase diagram of the proximity-induced gap in diffusive n-terminal Josephson junctions

    Science.gov (United States)

    Amundsen, Morten; Ouassou, Jabir Ali; Linder, Jacob

    2017-01-01

    Multiterminal Josephson junctions have recently been proposed as a route to artificially mimic topological matter with the distinct advantage that its properties can be controlled via the superconducting phase difference, giving rise to Weyl points in 4-terminal geometries. A key goal is to accurately determine when the system makes a transition from a gapped to non-gapped state as a function of the phase differences in the system, the latter effectively playing the role of quasiparticle momenta in conventional topological matter. We here determine the proximity gap phase diagram of diffusive n-terminal Josephson junctions (), both numerically and analytically, by identifying a class of solutions to the Usadel equation at zero energy in the full proximity effect regime. We present an analytical equation which provides the phase diagram for an arbitrary number of terminals n. After briefly demonstrating the validity of the analytical approach in the previously studied 2- and 3-terminal cases, we focus on the 4-terminal case and map out the regimes where the electronic excitations in the system are gapped and non-gapped, respectively, demonstrating also in this case full agreement between the analytical and numerical approach.

  11. Microprocessor, Setx, Xrn2, and Rrp6 co-operate to induce premature termination of transcription by RNAPII.

    Science.gov (United States)

    Wagschal, Alexandre; Rousset, Emilie; Basavarajaiah, Poornima; Contreras, Xavier; Harwig, Alex; Laurent-Chabalier, Sabine; Nakamura, Mirai; Chen, Xin; Zhang, Ke; Meziane, Oussama; Boyer, Frédéric; Parrinello, Hugues; Berkhout, Ben; Terzian, Christophe; Benkirane, Monsef; Kiernan, Rosemary

    2012-09-14

    Transcription elongation is increasingly recognized as an important mechanism of gene regulation. Here, we show that microprocessor controls gene expression in an RNAi-independent manner. Microprocessor orchestrates the recruitment of termination factors Setx and Xrn2, and the 3'-5' exoribonuclease, Rrp6, to initiate RNAPII pausing and premature termination at the HIV-1 promoter through cleavage of the stem-loop RNA, TAR. Rrp6 further processes the cleavage product, which generates a small RNA that is required to mediate potent transcriptional repression and chromatin remodeling at the HIV-1 promoter. Using chromatin immunoprecipitation coupled to high-throughput sequencing (ChIP-seq), we identified cellular gene targets whose transcription is modulated by microprocessor. Our study reveals RNAPII pausing and premature termination mediated by the co-operative activity of ribonucleases, Drosha/Dgcr8, Xrn2, and Rrp6, as a regulatory mechanism of RNAPII-dependent transcription elongation.

  12. Mitogen-activated protein kinases p38 and JNK mediate Actinobacillus pleuropneumoniae exotoxin ApxI-induced apoptosis in porcine alveolar macrophages.

    Science.gov (United States)

    Wu, Chi-Ming; Chen, Zeng-Weng; Chen, Ter-Hsin; Liao, Jiunn-Wang; Lin, Cheng-Chung; Chien, Maw-Sheng; Lee, Wei-Cheng; Hsuan, Shih-Ling

    2011-08-05

    Actinobacillus pleuropneumoniae exotoxins (Apx) are major virulence factors that play important roles in the pathogenesis of pleuropneumonia in swine. A previous study has demonstrated that native ApxI at low concentrations induces apoptosis in primary porcine alveolar macrophages (PAMs) via a caspase-3-dependent pathway. However, the molecular mechanisms underlying ApxI-induced apoptosis remain largely unknown. In this study, it was shown that ApxI treatment in PAMs rapidly induced phosphorylation of both p38 and JNK, members of the mitogen-activated protein kinase family. Application of a selective p38 or JNK inhibitor significantly reduced ApxI-induced apoptosis, indicating the involvement of p38 and JNK pathways in this event. Furthermore, activation of both caspase-8 and -9 were observed in ApxI-stimulated PAMs. Inhibition of caspase-8 and caspase-9 activity significantly protected PAMs from ApxI-induced apoptosis. In addition, Bid activation was also noted in ApxI-treated PAMs, and inhibition of caspase-8 suppressed the activation of Bid and caspase-9, suggesting that ApxI was able to activate the caspases-8-Bid-caspase-9 pathway. Notably, inhibition of p38 or JNK pathway greatly attenuated the activation of caspases-3, -8, and -9. This study is the first to demonstrate that ApxI-induced apoptosis of PAMs involves the activation of p38 and JNK, and engages the extrinsic and intrinsic apoptotic pathways.

  13. Study on human promyelocytic leukemia HL-60 cells apoptosis induced by fucosterol.

    Science.gov (United States)

    Ji, Yu-Bin; Ji, Chen-Feng; Yue, Lei

    2014-01-01

    In this study, we investigated the effect of fucosterol on HL-60 and the molecular mechanism. HL-60 Cells were treated with fucosterol, and 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) method was used to study fucosterol anti-tumor activity. Morphology of HL-60 cells was observed. Flow cytometry (FCM) was employed to detect the cell cycle. Laser scanning confocal microscope (LSCM) was used to analyze mitochondrial membrane potential (MMP) and the expressions of Fas, FasL, Fadd and Caspase-8. Western blot was performed to analyze the expressions of Cyt-C, Pro-Caspase-9 and Pro-Caspase-3. Caspase activity kits were used to determine the activity of Caspase-9, Caspase-8 and Caspase-3. The results showed fucosterol could inhibit the growth of HL-60 cells, and the cell cycle was arrested at G2/M phase. HL-60 cells showed obvious apoptosis morphology. After being treated with fucosterol for 24 h, HL-60 cells decreased MMP, induced Cyt-C release and Caspase-9, Caspase-3 activation. Fucosterol also increased the protein expression of Fas, FasL, Fadd and Caspase-8. Moreover, the activity of Caspase-9, Caspase-8 and Caspase-3 was increased significantly. In conclusion, Fucosterol can induce HL-60 cells apoptosis, suggesting that it may be a potent agent for cancer prevention and treatment.

  14. The Positively Charged COOH-terminal Glycosaminoglycan-binding CXCL9(74-103) Peptide Inhibits CXCL8-induced Neutrophil Extravasation and Monosodium Urate Crystal-induced Gout in Mice.

    Science.gov (United States)

    Vanheule, Vincent; Janssens, Rik; Boff, Daiane; Kitic, Nikola; Berghmans, Nele; Ronsse, Isabelle; Kungl, Andreas J; Amaral, Flavio Almeida; Teixeira, Mauro Martins; Van Damme, Jo; Proost, Paul; Mortier, Anneleen

    2015-08-28

    The ELR(-)CXC chemokine CXCL9 is characterized by a long, highly positively charged COOH-terminal region, absent in most other chemokines. Several natural leukocyte- and fibroblast-derived COOH-terminally truncated CXCL9 forms missing up to 30 amino acids were identified. To investigate the role of the COOH-terminal region of CXCL9, several COOH-terminal peptides were chemically synthesized. These peptides display high affinity for glycosaminoglycans (GAGs) and compete with functional intact chemokines for GAG binding, the longest peptide (CXCL9(74-103)) being the most potent. The COOH-terminal peptide CXCL9(74-103) does not signal through or act as an antagonist for CXCR3, the G protein-coupled CXCL9 receptor, and does not influence neutrophil chemotactic activity of CXCL8 in vitro. Based on the GAG binding data, an anti-inflammatory role for CXCL9(74-103) was further evidenced in vivo. Simultaneous intravenous injection of CXCL9(74-103) with CXCL8 injection in the joint diminished CXCL8-induced neutrophil extravasation. Analogously, monosodium urate crystal-induced neutrophil migration to the tibiofemural articulation, a murine model of gout, is highly reduced by intravenous injection of CXCL9(74-103). These data show that chemokine-derived peptides with high affinity for GAGs may be used as anti-inflammatory peptides; by competing with active chemokines for binding and immobilization on GAGs, these peptides may lower chemokine presentation on the endothelium and disrupt the generation of a chemokine gradient, thereby preventing a chemokine from properly performing its chemotactic function. The CXCL9 peptide may serve as a lead molecule for further development of inhibitors of inflammation based on interference with chemokine-GAG interactions.

  15. Stress-induced impairment of glutamatergic terminals ultrastructure: High vulnerability of medial prefrontal cortex and preventing action of desipramine

    DEFF Research Database (Denmark)

    Nava, N.; Popoli, M.; Musazzi, L.;

    2013-01-01

    mediators, glucocorticoids, on brain volume and dendritic remodeling, in both humans and rodents. Nevertheless, few is still known on the structural changes exerted by behavioral stress on the features of glutamatergic synapses as sites of neuronal communication. Indeed, in excitatory synapses synaptic...... communication is driven by neurotransmitter which is stored, within the presynaptic terminal, in morphologically distinct pools of vesicles, namely the readily-releasable pool of vesicles (RRP), docked to the active zone and ready for release, and the reserve pool of vesicles. When neurotransmitter is released...

  16. BRP, a polysaccharide fraction isolated from Boschniakia rossica, protects against galactosamine and lipopolysaccharide induced hepatic failure in mice.

    Science.gov (United States)

    Quan, Jishu; Jin, Meihua; Xu, Huixian; Qiu, Delai; Yin, Xuezhe

    2014-05-01

    The aim of this study was to investigate the hepatoprotective effect of BRP, a polysaccharide fraction isolated from Boschniakia rossica, against galactosamine and lipopolysaccharide induced fulminant hepatic failure. Mice were injected with a single dose of galactosamine/lipopolysaccharide with or without pretreatment of BRP. Results showed marked reduction of hepatic necrosis, serum marker enzymes and levels of tumor necrosis factor-α and interleukin-6 in BRP pretreated mice when compared with galactosamine/lipopolysaccharide-challenged mice. Mice pretreated with BRP decreased the activation of caspases-3 and caspase-8, and showed a reduced level of DNA fragmentation of liver cells. BRP also reduced hepatic lipid peroxidation, increased potential of hepatic antioxidative defense system, and reduced hepatic nitric oxide level which was elevated by galactosamine/lipopolysaccharide injection. Immunoblot analysis showed down-regulation of inducible nitric oxide synthase and cyclooxygenase-2 proteins of liver tissues in BRP pretreated group when compared with galactosamine/lipopolysaccharide-challenged group. Furthermore, treatment with galactosamine/lipopolysaccharide markedly increased toll-like receptor 4, nuclear level of nuclear factor-κB, and phosphorylation of both extracellular signal-regulated kinase and c-Jun N-terminal kinase in liver tissues. However, these increases were attenuated by pretreatment with BRP. The results suggest that BRP alleviates galactosamine/lipopolysaccharide-induced liver injury by enhancing antioxidative defense system, suppressing inflammatory responses and reducing apoptotic signaling.

  17. A C-Terminal Coiled-Coil Region of CagL is Responsible for Helicobacter Pylori-Induced Il-8 Expression.

    Science.gov (United States)

    Wiedemann, Tobias; Hofbaur, Stefan; Loell, Eva; Rieder, Gabriele

    2016-09-29

    Interleukin-8 (IL-8) is a potent neutrophil-activating chemokine which triggers the infiltration and migration of neutrophils into areas of bacterial infection. Helicobacter pylori-infected patient studies as well as animal models have revealed that H. pylori type I strains carrying an intact cytotoxin-associated gene pathogenicity island (cag-PAI) with a functional type IV secretion system (T4SS) induce IL-8 expression and secretion in gastric mucosa. This gastric mucosal IL-8 expression correlates with severe histological changes due to H. pylori infection. In the present study, we explored a new recognition pattern on the bacterial adhesion protein CagL inducing IL-8 expression in H. pylori-infected host cells. To analyze the secreted IL-8 concentration, we performed IL-8 enzyme-linked immunosorbent assay (ELISA). To investigate the H. pylori-induced IL-8 expression on the transcriptional level, we transiently transfected gastric epithelial cells (AGS) with a human IL-8 luciferase reporter construct. The results of this study demonstrate that specifically the C-terminal coiled-coil region of the H. pylori CagL protein, a protein described to be located on the tip of the T4SS-pilus, is responsible for several in vitro observations: 1) H. pylori-induced IL-8 secretion via the transforming growth factor (TGF)-α activated epidermal growth factor-receptor (EGF-R) signaling pathway; 2) H. pylori-induced elongation of the cells, a typical CagA-induced phenotype; and 3) the bridging of the T4SS to its human target cells. This novel bacterial-host recognition sequence allows a new insight into how H. pylori induces the inflammatory response in gastric epithelial cells and facilitates the development of precancerous conditions.

  18. A C-Terminal Coiled-Coil Region of CagL is Responsible for Helicobacter Pylori-Induced Il-8 Expression

    Science.gov (United States)

    Wiedemann, Tobias; Hofbaur, Stefan; Loell, Eva; Rieder, Gabriele

    2016-01-01

    Interleukin-8 (IL-8) is a potent neutrophil-activating chemokine which triggers the infiltration and migration of neutrophils into areas of bacterial infection. Helicobacter pylori-infected patient studies as well as animal models have revealed that H. pylori type I strains carrying an intact cytotoxin-associated gene pathogenicity island (cag-PAI) with a functional type IV secretion system (T4SS) induce IL-8 expression and secretion in gastric mucosa. This gastric mucosal IL-8 expression correlates with severe histological changes due to H. pylori infection. In the present study, we explored a new recognition pattern on the bacterial adhesion protein CagL inducing IL-8 expression in H. pylori-infected host cells. To analyze the secreted IL-8 concentration, we performed IL-8 enzyme-linked immunosorbent assay (ELISA). To investigate the H. pylori-induced IL-8 expression on the transcriptional level, we transiently transfected gastric epithelial cells (AGS) with a human IL-8 luciferase reporter construct. The results of this study demonstrate that specifically the C-terminal coiled-coil region of the H. pylori CagL protein, a protein described to be located on the tip of the T4SS-pilus, is responsible for several in vitro observations: 1) H. pylori-induced IL-8 secretion via the transforming growth factor (TGF)-α activated epidermal growth factor-receptor (EGF-R) signaling pathway; 2) H. pylori-induced elongation of the cells, a typical CagA-induced phenotype; and 3) the bridging of the T4SS to its human target cells. This novel bacterial-host recognition sequence allows a new insight into how H. pylori induces the inflammatory response in gastric epithelial cells and facilitates the development of precancerous conditions. PMID:27766167

  19. Infection of epithelial cells with Chlamydia trachomatis inhibits TNF-induced apoptosis at the level of receptor internalization while leaving non-apoptotic TNF-signalling intact.

    Science.gov (United States)

    Waguia Kontchou, Collins; Tzivelekidis, Tina; Gentle, Ian E; Häcker, Georg

    2016-11-01

    Chlamydia trachomatis is an obligate intracellular bacterial pathogen of medical importance. C. trachomatis develops inside a membranous vacuole in the cytosol of epithelial cells but manipulates the host cell in numerous ways. One prominent effect of chlamydial infection is the inhibition of apoptosis in the host cell, but molecular aspects of this inhibition are unclear. Tumour necrosis factor (TNF) is a cytokine with important roles in immunity, which is produced by immune cells in chlamydial infection and which can have pro-apoptotic and non-apoptotic signalling activity. We here analysed the signalling through TNF in cells infected with C. trachomatis. The pro-apoptotic signal of TNF involves the activation of caspase-8 and is controlled by inhibitor of apoptosis proteins. We found that in C. trachomatis-infected cells, TNF-induced apoptosis was blocked upstream of caspase-8 activation even when inhibitor of apoptosis proteins were inhibited or the inhibitor of caspase-8 activation, cFLIP, was targeted by RNAi. However, when caspase-8 was directly activated by experimental over-expression of its upstream adapter Fas-associated protein with death domain, C. trachomatis was unable to inhibit apoptosis. Non-apoptotic TNF-signalling, particularly the activation of NF-κB, initiates at the plasma membrane, while the activation of caspase-8 and pro-apoptotic signalling occur subsequently to internalization of TNF receptor and the formation of a cytosolic signalling complex. In C. trachomatis-infected cells, NF-κB activation through TNF was unaffected, while the internalization of the TNF-TNF-receptor complex was blocked, explaining the lack of caspase-8 activation. These results identify a dichotomy of TNF signalling in C. trachomatis-infected cells: Apoptosis is blocked at the internalization of the TNF receptor, but non-apoptotic signalling through this receptor remains intact, permitting a response to this cytokine at sites of infection.

  20. Calcium absorption by Cav1.3 induces terminal web myosin II phosphorylation and apical GLUT2 insertion in rat intestine.

    Science.gov (United States)

    Mace, Oliver J; Morgan, Emma L; Affleck, Julie A; Lister, Norma; Kellett, George L

    2007-04-15

    Glucose absorption in rat jejunum involves Ca(2+)- and PKC betaII-dependent insertion of GLUT2 into the apical membrane. Ca(2+)-induced rearrangement of the enterocyte cytoskeleton is thought to enhance paracellular flow. We have therefore investigated the relationships between myosin II regulatory light chain phosphorylation (RLC(20)), absorption of glucose, water and calcium, and mannitol clearance. ML-7, an inhibitor of myosin light chain kinase, diminished the phloretin-sensitive apical GLUT2 but not the phloretin-insensitive SGLT1 component of glucose absorption in rat jejunum perfused with 75 mM glucose. Western blotting and immunocytochemistry revealed marked decreases in RLC(20) phosphorylation in the terminal web and in the levels of apical GLUT2 and PKC betaII, but not SGLT1. Perfusion with phloridzin or 75 mM mannitol, removal of luminal Ca(2+), or inhibition of unidirectional (45)Ca(2+) absorption by nifedipine exerted similar effects. ML-7 had no effect on the absorption of 10 mM Ca(2+), nor clearance of [(14)C]-mannitol, which was less than 0.7% of the rate of glucose absorption. Water absorption did not correlate with (45)Ca(2+) absorption or mannitol clearance. We conclude that the Ca(2+) necessary for contraction of myosin II in the terminal web enters via an L-type channel, most likely Ca(v)1.3, and is dependent on SGLT1. Moreover, terminal web RLC(20) phosphorylation is necessary for apical GLUT2 insertion. The data confirm that glucose absorption by paracellular flow is negligible, and show further that paracellular flow makes no more than a minimal contribution to jejunal Ca(2+) absorption at luminal concentrations prevailing after a meal.

  1. Intrathecal administration of low-dose nociceptin/orphanin FQ induces allodynia via c-Jun N-terminal kinase and monocyte chemoattractant protein-1.

    Science.gov (United States)

    Kawabata, Kenta; Nishimura, Isamu; Fujiwara, Takeshi; Terauchi, Shoko; Minami, Toshiaki; Ito, Seiji; Okuda-Ashitaka, Emiko

    2016-06-01

    Pathological chronic pain, which is frequently associated with prolonged tissue damage, inflammation, tumour invasion, and neurodegenerative diseases, gives rise to hyperalgesia and allodynia. We previously reported that intrathecal administration of nociceptin/orphanin FQ (N/OFQ), an endogenous ligand for the orphan opioid receptor-like receptor, in the femtomole range induces touch-evoked allodynia. N/OFQ has been implicated in multiple signalling pathways, such as inhibition of cAMP production and Ca(2+) channels, or activation of K(+) channels and mitogen-activated protein kinase, although the signalling pathways of N/OFQ-induced allodynia remain unclear. To address these issues, we developed an ex vivo mitogen-activated protein kinase assay by using intact slices of mouse spinal cord. N/OFQ markedly increased the phosphorylation of c-Jun N-terminal kinase (JNK) in the superficial dorsal horn of the spinal cord. The N/OFQ-stimulated JNK phosphorylation was significantly inhibited by pertussis toxin, the phospholipase C inhibitor U73122, and the inositol trisphosphate receptor antagonist Xestospongin C. Intrathecal administration of the JNK inhibitor SP600125 inhibited N/OFQ-evoked allodynia. The N/OFQ-induced increase in JNK phosphorylation was observed in astrocytes that expressed glial fibrillary acidic protein. N/OFQ also induced monocyte chemoattractant protein-1 (MCP-1) release via the JNK pathway, and N/OFQ-induced JNK phosphorylation was observed in MCP-1-immunoreactive astrocytes. Intrathecal administration of the MCP-1 receptor antagonist RS504393 inhibited N/OFQ-evoked allodynia. These results suggest that, in the spinal dorsal horn, N/OFQ induces allodynia through activation of JNK via the phospholipase C-inositol trisphosphate pathway, which is coupled to pertussis toxin-sensitive G-protein, and following the release of MCP-1 from astrocytes.

  2. N-(1-Pyrenyl Maleimide Induces Bak Oligomerization and Mitochondrial Dysfunction in Jurkat Cells

    Directory of Open Access Journals (Sweden)

    Pei-Rong Huang

    2015-01-01

    Full Text Available N-(1-pyrenyl maleimide (NPM is a fluorescent reagent that is frequently used as a derivatization agent for the detection of thio-containing compounds. NPM has been shown to display a great differential cytotoxicity against hematopoietic cancer cells. In this study, the molecular mechanism by which NPM induces apoptosis was examined. Here, we show that treatment of Jurkat cells with NPM leads to Bak oligomerization, loss of mitochondrial membrane potential (Δψm, and release of cytochrome C from mitochondria to cytosol. Induction of Bak oligomerization appears to play a critical role in NPM-induced apoptosis, as downregulation of Bak by shRNA significantly prevented NPM-induced apoptosis. Inhibition of caspase 8 by Z-IETD-FMK and/or depletion of Bid did not affect NPM-induced oligomerization of Bak. Taken together, these results suggest that NPM-induced apoptosis is mediated through a pathway that is independent of caspase-8 activation.

  3. Depletion of histone N-terminal-acetyltransferase Naa40 induces p53-independent apoptosis in colorectal cancer cells via the mitochondrial pathway.

    Science.gov (United States)

    Pavlou, Demetria; Kirmizis, Antonis

    2016-03-01

    Protein N-terminal acetylation is an abundant post-translational modification in eukaryotes implicated in various fundamental cellular and biochemical processes. This modification is catalysed by evolutionarily conserved N-terminal acetyltransferases (NATs) whose deregulation has been linked to cancer development and thus, are emerging as useful diagnostic and therapeutic targets. Naa40 is a highly selective NAT that acetylates the amino-termini of histones H4 and H2A and acts as a sensor of cell growth in yeast. In the present study, we examine the role of Naa40 in cancer cell survival. We demonstrate that depletion of Naa40 in HCT116 and HT-29 colorectal cancer cells decreases cell survival by enhancing apoptosis, whereas Naa40 reduction in non-cancerous mouse embryonic fibroblasts has no effect on cell viability. Specifically, Naa40 knockdown in colon cancer cells activates the mitochondrial caspase-9-mediated apoptotic cascade. Consistent with this, we show that caspase-9 activation is required for the induced apoptosis because treatment of cells with an irreversible caspase-9 inhibitor impedes apoptosis when Naa40 is depleted. Furthermore, the effect of Naa40-depletion on cell-death is mediated through a p53-independent mechanism since p53-null HCT116 cells still undergo apoptosis upon reduction of the acetyltransferase. Altogether, these findings reveal an anti-apoptotic role for Naa40 and exhibit its potential as a therapeutic target in colorectal cancers.

  4. Terminal structure

    Science.gov (United States)

    Schmidt, Frank; Allais, Arnaud; Mirebeau, Pierre; Ganhungu, Francois; Lallouet, Nicolas

    2009-10-20

    A terminal structure (2) for a superconducting cable (1) is described. It consists of a conductor (2a) and an insulator (2b) that surrounds the conductor (2a), wherein the superconducting cable (1) has a core with a superconducting conductor (5) and a layer of insulation that surrounds the conductor (5), and wherein the core is arranged in such a way that it can move longitudinally in a cryostat. The conductor (2a) of the terminal structure (2) is electrically connected with the superconducting conductor (5) or with a normal conductor (6) that is connected with the superconducting conductor (5) by means of a tubular part (7) made of an electrically conductive material, wherein the superconducting conductor (5) or the normal conductor (6) can slide in the part (7) in the direction of the superconductor.

  5. The cathepsin B inhibitor, z-FA-CMK is toxic and readily induced cell death in human T lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Liow, K.Y.; Chow, S.C., E-mail: chow.sek.chuen@monash.edu

    2013-11-01

    The cathepsin B inhibitor, benzyloxycarbonyl-phenylalanine-alanine-chloromethylketone (z-FA-CMK) was found to be toxic and readily induced cell death in the human T cell line, Jurkat, whereas two other analogs benzyloxycarbonyl-phenylalanine-alanine-fluoromethylketone (z-FA-FMK) and benzyloxycarbonyl-phenylalanine-alanine-diazomethylketone (z-FA-DMK) were not toxic. The toxicity of z-FA-CMK requires not only the CMK group, but also the presence of alanine in the P1 position and the benzyloxycarbonyl group at the N-terminal. Dose–response studies showed that lower concentrations of z-FA-CMK induced apoptosis in Jurkat T cells whereas higher concentrations induced necrosis. In z-FA-CMK-induced apoptosis, both initiator caspases (-8 and -9) and effector caspases (-3, -6 and -7) were processed to their respective subunits in Jurkat T cells. However, only the pro-form of the initiator caspases were reduced in z-FA-CMK-induced necrosis and no respective subunits were apparent. The caspase inihibitor benzyloxycarbonyl-valine-alanine-aspartic acid-(O-methyl)-fluoromehylketone (z-VAD-FMK) inhibits apoptosis and caspase processing in Jurkat T cells treated with low concentration of z-FA-CMK but has no effect on z-FA-CMK-induced necrosis and the loss of initiator caspases. This suggests that the loss of initiator caspases in Jurkat T cells during z-FA-CMK-induced necrosis is not a caspase-dependent process. Taken together, we have demonstrated that z-FA-CMK is toxic to Jurkat T cells and induces apoptosis at low concentrations, while at higher concentrations the cells die of necrosis. - Highlights: • z-FA-CMK is toxic and induce cell death in the human T cells. • z-FA-CMK toxicity requires the CMK group, alanine and the benzyloxycarbonyl group. • z-FA-CMK induced apoptosis at low concentration and necrosis at high concentration.

  6. Termination unit

    Energy Technology Data Exchange (ETDEWEB)

    Traeholt, Chresten; Willen, Dag; Roden, Mark; Tolbert, Jerry C.; Lindsay, David; Fisher, Paul W.; Nielsen, Carsten Thidemann

    2016-05-03

    Cable end section comprises end-parts of N electrical phases/neutral, and a thermally-insulation envelope comprising cooling fluid. The end-parts each comprises a conductor and are arranged with phase 1 innermost, N outermost surrounded by the neutral, electrical insulation being between phases and N and neutral. The end-parts comprise contacting surfaces located sequentially along the longitudinal extension of the end-section. A termination unit has an insulating envelope connected to a cryostat, special parts at both ends comprising an adapter piece at the cable interface and a closing end-piece terminating the envelope in the end-section. The special parts houses an inlet and/or outlet for cooling fluid. The space between an inner wall of the envelope and a central opening of the cable is filled with cooling fluid. The special part at the end connecting to the cryostat houses an inlet or outlet, splitting cooling flow into cable annular flow and termination annular flow.

  7. (-)-Bornyl acetate induces autonomic relaxation and reduces arousal level after visual display terminal work without any influences of task performance in low-dose condition.

    Science.gov (United States)

    Matsubara, Eri; Fukagawa, Mio; Okamoto, Tsuyoshi; Ohnuki, Koichiro; Shimizu, Kuniyoshi; Kondo, Ryuichiro

    2011-04-01

    (-)-Bornyl acetate is the main volatile constituent in numerous conifer oils and has a camphoraceous, pine-needle-like odor. It is frequently used as the conifer needle composition in soap, bath products, room sprays, and pharmaceutical products. However, the psychophysiological effects of (-)-bornyl acetate remained unclear. We investigated the effects of breathing air mixed with (-)-bornyl acetate at different doses (low-dose and high-dose conditions) on the individuals during and after VDT (visual display terminal) work using a visual discrimination task. The amounts of (-)-bornyl acetate through our odorant delivery system for 40 min were 279.4 µg in the low-dose and 716.3 µg in the high-dose (-)-bornyl acetate condition. (-)-Bornyl acetate induced changes of autonomic nervous system for relaxation and reduced arousal level after VDT work without any influences of task performance in low-dose condition, but not in high-dose condition.

  8. Role of N-terminal residues on folding and stability of C-phycoerythrin: simulation and urea-induced denaturation studies.

    Science.gov (United States)

    Anwer, Khalid; Sonani, Ravi; Madamwar, Datta; Singh, Parvesh; Khan, Faez; Bisetty, Krishna; Ahmad, Faizan; Hassan, Md Imtaiyaz

    2015-01-01

    The conformational state of biliproteins can be determined by optical properties of the covalently linked chromophores. Recently determined crystal structure of truncated form of α-subunit of cyanobacterial phycoerythrin (αC-PE) from Phormidium tenue provides a new insight into the structure-function relationship of αC-PE. To compare their stabilities, we have measured urea-induced denaturation transitions of the full length αC-PE (FL-αC-PE) and truncated αC-PE (Tr-αC-PE) followed by observing changes in absorbance at 565 nm, fluorescence at 350 and 573 nm, and circular dichroism at 222 nm as a function of [urea], the molar concentration of urea. The transition curve of each protein was analyzed for ΔG(D)(0), the value of Gibbs free energy change on denaturation (ΔG(D)) in the absence of urea; m, the slope (=∂∆G(D)/∂[urea]), and C(m), the midpoint of the denaturation curve, i.e. [urea] at which ΔG(D) = 0. A difference of about 10% in ΔG(D)(0) observed between FL-αC-PE and Tr-αC-PE, suggests that the two proteins are almost equally stable, and the natural deletion of 31 residues from the N-terminal side of the full length protein does not alter its stability. Furthermore, normalization of probes shows that the urea-induced denaturation of both the proteins is a two-state process. Folding of both structural variants (Tr-αC-PE and FL-αC-PE) of P. tenue were also studied using molecular dynamics simulations at 300 K. The results show clearly that the stability of the proteins is evenly distributed over the whole structure indicating no significant role of N-terminal residues in the stability of both proteins.

  9. Pseudomonas aeruginosa Ventilator-Associated Pneumonia Induces Lung Injury through TNF-α/c-Jun NH2-Terminal Kinase Pathways

    Science.gov (United States)

    Hsu, Ching-Mei

    2017-01-01

    Ventilator-associated pneumonia (VAP) is a common nosocomial infection among intensive care unit (ICU) patients. Pseudomonas aeruginosa (PA) is the most common multidrug-resistant Gram-negative pathogen and VAP caused by PA carries a high rate of morbidity and mortality. This study examined the molecular mechanism of PA VAP-induced lung injury. C57BL/6 wild-type (WT) mice and JNK1 knockout (JNK1-/-) mice received mechanical ventilation (MV) for 3 h at 2 days after receiving nasal instillation of PA. The WT and JNK1-/- mice also received MV after the induction of lung injury by instillation of supernatants from PA-stimulated alveolar macrophages (AMs). AMs isolated from WT, IκB-kinase (IKK)βΔMye (IKKβ was selectively deleted in macrophages), and JNK1-/- mice were ex vivo stimulated with live PA and supernatants were collected for cytokine assay. Intranasal instillation of 106 PA enhanced MV-induced NF-κB DNA binding activity in the lungs and nitrite levels in BALF. MV after PA instillation significantly increased the expression of ICAM and VCAM in the lungs and TNF-α, IL-1β, and IL-6 levels in bronchoalveolar lavage fluid (BALF) of WT mice, but not in JNK1-/- mice. MV after supernatant instillation induced more total protein concentration in BALF and neutrophil sequestration in the lungs in WT mice than JNK1-/- mice and cytokine assay of supernatants indicated that TNF-α is a critical regulator of PA VAP-induced lung injury. Ex vivo PA stimulation induced TNF-α production by AMs from WT as well as JNK1-/- mice but not IKKβΔMye mice. In summary, PA colonization plays an important role in PA VAP-induced lung injury through the induction of JNK1-mediated inflammation. These results suggest that the pathogenesis mechanism of PA VAP involves production of TNF-α through activation of IKK/NF-κB pathways in AMs and JNK signaling pathway in the lungs. PMID:28060857

  10. TJ-41 Induces Apoptosis and Potentiates the Apoptotic Effects of 5-FU in Breast Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Suresh Volate

    2009-01-01

    Full Text Available Recent studies suggest that TJ-41, a herbal drug, possesses chemotherapeutic effects. Accordingly, this study was undertaken to investigate the anticarcinogenic effects of TJ-41 on human breast cancer cells lines. TJ-41 inhibited the proliferation of human breast cancer cell lines dose dependently. Flow cytometric analysis showed that this decrease in DNA synthesis is to be associated with induction of apoptosis. In both cell lines, apoptosis was abolished by caspase-9 inhibitor Z-LEHD-fmk but was weakly inhibited by caspase-8 inhibitor Z-IETD-fmk, indicating that caspase-9 activation was involved in TJ-41 induced apoptosis. Additionally, TJ-41 stimulated phosphorylation of c-Jun NH2-terminal kinase (JNK and pretreatment of breast cancer cells with JNK inhibitor SP600125 completely abolished TJ-41 induced apoptosis. Our data also demonstrate that combined treatment of TJ-41 and 5-FU significantly potentiates the apoptotic effects of 5-FU in both breast cancer cell lines. Taken together, these data suggest that TJ-41 might provide a novel chemotherapeutic treatment for breast cancer.

  11. (-)-Epigallocatechin-3-gallate decreases thrombin/paclitaxel-induced endothelial tissue factor expression via the inhibition of c-Jun terminal NH2 kinase phosphorylation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Huang-Joe [Institute of Biotechnology, National Tsing Hua University, No. 101, Section 2, Kuang Fu Road, Hsinchu 30013, Taiwan (China); Division of Cardiology, Department of Medicine, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung 40447, Taiwan (China); Lo, Wan-Yu [Department of Medical Research, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung 40447, Taiwan (China); Graduate Integration of Chinese and Western Medicine, China Medical University, No. 91, Hsueh-Shih Road, Taichung 40402, Taiwan (China); Lu, Te-Ling [School of Pharmacy, China Medical University, No. 91, Hsueh-Shih Road, Taichung 40402, Taiwan (China); Huang, Haimei, E-mail: hmhuang@life.nthu.edu.tw [Institute of Biotechnology, National Tsing Hua University, No. 101, Section 2, Kuang Fu Road, Hsinchu 30013, Taiwan (China)

    2010-01-01

    Patients with paclitaxel-eluting stents are concerned with stent thrombosis caused by premature discontinuation of dual antiplatelet therapy or clopidogrel resistance. This study investigates the effect of (-)-epigallocatechin-3-gallate (EGCG) on the expression of thrombin/paclitaxel-induced endothelial tissue factor (TF) expressions in human aortic endothelial cells (HAECs). EGCG was nontoxic to HAECs at 6 h up to a concentration of 25 {mu}mol/L. At a concentration of 25 {mu}mol/L, EGCG pretreatment potently inhibited both thrombin-stimulated and thrombin/paclitaxel-stimulated endothelial TF protein expression. Thrombin and thrombin/paclitaxel-induced 2.6-fold and 2.9-fold increases in TF activity compared with the control. EGCG pretreatment caused a 29% and 38% decrease in TF activity on thrombin and thrombin/paclitaxel treatment, respectively. Real-time polymerase chain reaction (PCR) showed that thrombin and thrombin/paclitaxel-induced 3.0-fold and 4.6-fold TF mRNA expressions compared with the control. EGCG pretreatment caused an 82% and 72% decrease in TF mRNA expression on thrombin and thrombin/paclitaxel treatment, respectively. The c-Jun terminal NH2 kinase (JNK) inhibitor SP600125 reduced thrombin/paclitaxel-induced TF expression. Furthermore, EGCG significantly inhibited the phosphorylation of JNK to 49% of thrombin/paclitaxel-stimulated HAECs at 60 min. Immunofluorescence assay did not show an inhibitory effect of EGCG on P65 NF-{kappa}B nuclear translocation in the thrombin/paclitaxel-stimulated endothelial cells. In conclusion, EGCG can inhibit TF expression in thrombin/paclitaxel-stimulated endothelial cells via the inhibition of JNK phosphorylation. The unique property of EGCG may be used to develop a new drug-eluting stent by co-coating EGCG and paclitaxel.

  12. S-adenosyl-methionine decreases ethanol-induced apoptosis in primary hepatocyte cultures by a c-Jun N-terminal kinase activity-independent mechanism

    Institute of Scientific and Technical Information of China (English)

    María del Pilar Cabrales-Romero; Lucrecia Márquez-Rosado; Samia Fattel-Fazenda; Cristina Trejo-Solís; Evelia Arce-Popoca; Leticia Alemén-Lazarini; Saúl Villa-Trevi(n)o

    2006-01-01

    AIM: To determine the role of c-Jun N-terminal kinase (JNK) activity in ethanol-induced apoptosis and the modulation of this signaling cascade by S-Adenosylmethionine (AdoMet).METHODS: Primary hepatocyte cultures were pretreated with 100 μmol/L SP600125, a selective JNK inhibitor, 1 mL/L DMSO or 4 mmol/L AdoMet and then exposed to 100 mmo/L ethanol. Hepatocyte apoptosis was determined by the TUNEL and DNA ladder assays.JNK activity and its inhibition by SP600125 and AdoMet were determined by Western blot analysis of c-jun phosphorylation and Bid fragmentation. SP600125 and AdoMet effects on the apoptotic signaling pathway were determined by Western blot analysis of cytochrome c release and pro-caspase 3 fragmentation. The AdoMet effect on glutathione levels was measured by Ellman's method and reactive oxygen species (ROS) generation by cell cytometry.RESULTS: The exposure of hepatocytes to ethanol induced JNK activation, c-jun phosphorylation, Bid fragmentation, cytochrome c release and pro-caspase 3 cleavage; these effects were diminished by SP600125, and caused a significant decreasein ethanol-induced apoptosis (P< 0.05). AdoMet exerted an antioxidant effect maintaining glutathione levels and decreasing ROS generation, without a significant effect on JNK activity,and prevented cytochrome c release and pro-caspase 3 cleavage.CONCLUSION: The JNK signaling cascade is a key component of the proapoptotic signaling pathway induced by ethanol. JNK activation may be independent from ROS generation, since AdoMet which exerted antioxidant properties did not have a significant effect on JNK activity. JNK pathway modulator agents and AdoMet may be components of promising therapies for alcoholic liver disease (ALD) treatment.

  13. c-Jun N-terminal kinase is required for vitamin E succinate-induced apoptosis in human gastric cancer cells

    Institute of Scientific and Technical Information of China (English)

    Kun Wu; Yan Zhao; Gui-Chang Li; Wei-Ping Yu

    2004-01-01

    AIM: To investigate the roles of c-Jun N-terminal kinase (JNK)signaling pathway in vitamin E succinate-induced apoptosis in human gastric cancer SGC-7901 cells.METHODS: Human gastric cancer cell lines (SGC-7901)were treated with vitamin E succinate (VES) at 5, 10, 20 mg/L.Succinic acid and vitamin E were used as vehicle controls and condition medium only as an untreated (UT) control.Apoptosis was observed by 4′, 6-diamidine-2′-phenylindole dihydrochloride (DAPI) staining for morphological changes and by DNA fragmentation for biochemical alterations.Western blot analysis was applied to measure the expression ofJNK and phosphorylated JNK. After the cells were transiently transfected with dominant negative mutant of JNK (DNJNK) followed by treatment of VES, the expression of JNK and c-Jun protein was determined.RESULTS: The apoptotic changes were observed after VES treatment by DNA fragmentation. DNA ladder in the 20 mg/L VES group was more clearly seen than that in 10 mg/L VES group and was not detected following treatment of UT control, succinate and vitamin E. VES at 5, 10 and 20 mg/L increased the expression of p-JNK by 2.5-, 2.8- and 4.2-fold, respectively. VES induced the phosphorylation of JNK beginning at 1.5 h and produced a sustained increase for 24 h with the peak level at 12 h. Transient transfection of DN-JNK blocked VES-triggered apoptosis by 52%. DN-JNK significantly increased the level of JNK, while decreasing the expression of VES-induced c-Jun protein.CONCLUSION: VES-induced apoptosis in human gastric cancer SGC-7901 cells involves JNK signaling pathway via c-Jun and its downstream transcription factor.

  14. Glucose, Lactate and Glutamine but not Glutamate Support Depolarization-Induced Increased Respiration in Isolated Nerve Terminals

    DEFF Research Database (Denmark)

    Hohnholt, Michaela C; Andersen, Vibe H; Bak, Lasse K;

    2016-01-01

    and glycolytic rate in presence of different substrates. Mitochondrial function was tested by sequentially exposure of the synaptosomes to the ATP synthase inhibitor, oligomycin, the uncoupler FCCP (carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone) and the electron transport chain inhibitors rotenone...... and antimycin A. The synaptosomes exhibited intense respiratory activity using glucose as substrate. The FCCP-dependent respiration was significantly higher with 10 mM glucose compared to 1 mM glucose. Synaptosomes also readily used pyruvate as substrate, which elevated basal respiration, activity....... Synaptosomal respiration using glutamate and glutamine as substrates was significantly higher compared to basal respiration, whereas oligomycin-dependent and FCCP-induced respiration was lower compared to the responses obtained in the presence of glucose as substrate. We provide evidence that synaptosomes...

  15. Glucose, Lactate and Glutamine but not Glutamate Support Depolarization-Induced Increased Respiration in Isolated Nerve Terminals.

    Science.gov (United States)

    Hohnholt, Michaela C; Andersen, Vibe H; Bak, Lasse K; Waagepetersen, Helle S

    2017-01-01

    Synaptosomes prepared from various aged and gene modified experimental animals constitute a valuable model system to study pre-synaptic mechanisms. Synaptosomes were isolated from whole brain and the XFe96 extracellular flux analyzer (Seahorse Bioscience) was used to study mitochondrial respiration and glycolytic rate in presence of different substrates. Mitochondrial function was tested by sequentially exposure of the synaptosomes to the ATP synthase inhibitor, oligomycin, the uncoupler FCCP (carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone) and the electron transport chain inhibitors rotenone and antimycin A. The synaptosomes exhibited intense respiratory activity using glucose as substrate. The FCCP-dependent respiration was significantly higher with 10 mM glucose compared to 1 mM glucose. Synaptosomes also readily used pyruvate as substrate, which elevated basal respiration, activity-dependent respiration induced by veratridine and the respiratory response to uncoupling compared to that obtained with glucose as substrate. Also lactate was used as substrate by synaptosomes but in contrast to pyruvate, mitochondrial lactate mediated respiration was comparable to respiration using glucose as substrate. Synaptosomal respiration using glutamate and glutamine as substrates was significantly higher compared to basal respiration, whereas oligomycin-dependent and FCCP-induced respiration was lower compared to the responses obtained in the presence of glucose as substrate. We provide evidence that synaptosomes are able to use besides glucose and pyruvate also the substrates lactate, glutamate and glutamine to support their basal respiration. Veratridine was found to increase respiration supported by glucose, pyruvate, lactate and glutamine and FCCP was found to increase respiration supported by glucose, pyruvate and lactate. This was not the case when glutamate was the only energy substrate.

  16. Midazolam induces apoptosis in MA-10 mouse Leydig tumor cells through caspase activation and the involvement of MAPK signaling pathway

    Directory of Open Access Journals (Sweden)

    So EC

    2014-02-01

    Full Text Available Edmund Cheung So,1,2 Yu-Xuan Lin,3 Chi Hao Tseng,1 Bo-Syong Pan,3 Ka-Shun Cheng,2 Kar-Lok Wong,2 Lyh-Jyh Hao,4 Yang-Kao Wang,5 Bu-Miin Huang2 1Department of Anesthesia, Tainan Municipal An Nan Hospital, China Medical University, Tainan, Taiwan; 2Department of Anesthesia, China Medical University, Taichung, Taiwan; 3Department of Cell Biology and Anatomy, National Cheng Kung University, Tainan, Taiwan; 4Department of Internal Medicine, Division of Endocrinology and Metabolism, Kaohsiung Veteran General Hospital Tainan Branch Tainan, Taiwan; 5Graduate Institute of Biomedical Materials and Tissue Engineering, Taipei Medical University, Taipei, Taiwan Purpose: The present study aims to investigate how midazolam, a sedative drug for clinical use with cytotoxicity on neuronal and peripheral tissues, induced apoptosis in MA-10 mouse Leydig tumor cells. Methods: The apoptotic effect and underlying mechanism of midazolam to MA-10 cells were investigated by flow cytometry assay and Western blotting methods. Results: Data showed that midazolam induced the accumulation of the MA-10 cell population in the sub-G1 phase and a reduction in the G2/M phase in a time- and dose-dependent manner, suggesting an apoptotic phenomenon. Midazolam could also induce the activation of caspase-8, -9, and -3 and poly (ADP-ribose polymerase proteins. There were no changes in the levels of Bax and cytochrome-c, whereas Bid was significantly decreased after midazolam treatment. Moreover, midazolam decreased both pAkt and Akt expression. In addition, midazolam stimulated the phosphorylation of p38 and c-Jun NH2-terminal kinase but not extracellular signal-regulated kinase. Conclusion: Midazolam could induce MA-10 cell apoptosis through the activation of caspase cascade, the inhibition of pAkt pathway, and the induction of p38 and c-Jun NH2-terminal kinase pathways. Keywords: midazolam, apoptosis, MA-10 cell, caspase, Akt, MAPKs

  17. Streptococcus pneumoniae induced c-Jun-N-terminal kinase- and AP-1 -dependent IL-8 release by lung epithelial BEAS-2B cells

    Directory of Open Access Journals (Sweden)

    Rosseau Simone

    2006-07-01

    Full Text Available Abstract Background Although pneumococcal pneumonia is one of the most common causes of death due to infectious diseases, little is known about pneumococci-lung cell interaction. Herein we tested the hypothesis that pneumococci activated pulmonary epithelial cell cytokine release by c-Jun-NH2-terminal kinase (JNK Methods Human bronchial epithelial cells (BEAS-2B or epithelial HEK293 cells were infected with S. pneumoniae R6x and cytokine induction was measured by RT-PCR, ELISA and Bioplex assay. JNK-phosphorylation was detected by Western blot and nuclear signaling was assessed by electrophoretic mobility shift assay (EMSA and chromatin immunoprecipitation (ChIP. JNK was modulated by the small molecule inhibitor SP600125 and AP1 by transfection of a dominant negative mutant. Results S. pneumoniae induced the release of distinct CC and CXC, as well as Th1 and Th2 cytokines and growth factors by human lung epithelial cell line BEAS-2B. Furthermore, pneumococci infection resulted in JNK phosphorylation in BEAS-2B cells. Inhibition of JNK by small molecule inhibitor SP600125 reduced pneumococci-induced IL-8 mRNA expression and release of IL-8 and IL-6. One regulator of the il8 promoter is JNK-phosphorylated activator protein 1 (AP-1. We showed that S. pneumoniae time-dependently induced DNA binding of AP-1 and its phosphorylated subunit c-Jun with a maximum at 3 to 5 h after infection. Recruitment of Ser63/73-phosphorylated c-Jun and RNA polymerase II to the endogenous il8 promoter was found 2 h after S. pneumoniae infection by chromatin immunoprecipitation. AP-1 repressor A-Fos reduced IL-8 release by TLR2-overexpressing HEK293 cells induced by pneumococci but not by TNFα. Antisense-constructs targeting the AP-1 subunits Fra1 and Fra2 had no inhibitory effect on pneumococci-induced IL-8 release. Conclusion S. pneumoniae-induced IL-8 expression by human epithelial BEAS-2B cells depended on activation of JNK and recruitment of phosphorylated c

  18. Terminal ballistics

    CERN Document Server

    Rosenberg, Zvi

    2016-01-01

    This book comprehensively discusses essential aspects of terminal ballistics, combining experimental data, numerical simulations and analytical modeling. Employing a unique approach to numerical simulations as a measure of sensitivity for the major physical parameters, the new edition also includes the following features: new figures to better illustrate the problems discussed; improved explanations for the equation of state of a solid and for the cavity expansion process; new data concerning the Kolsky bar test; and a discussion of analytical modeling for the hole diameter in a thin metallic plate impacted by a shaped charge jet. The section on thick concrete targets penetrated by rigid projectiles has now been expanded to include the latest findings, and two new sections have been added: one on a novel approach to the perforation of thin concrete slabs, and one on testing the failure of thin metallic plates using a hydrodynamic ram.

  19. Ligand-controlled interaction of histone acetyltransferase binding to ORC-1 (HBO1) with the N-terminal transactivating domain of progesterone receptor induces steroid receptor coactivator 1-dependent coactivation of transcription

    NARCIS (Netherlands)

    M. Georgiakaki (Maria); L.J. Blok (Leen); R. Milgrom (Roni); M. Lombès (Marc); A. Guiochon-Mantel (Anne); H. Loosfelt (Hugues); N. Chabbert-Buffet (Nathalie); B. Dasen (Boris); G. Meduri (Geri); S. Wenk (Sandra); L. Rajhi (Leila); L. Amazit (Larbi); A. Chauchereau (Anne); C.W. Burger (Curt)

    2006-01-01

    textabstractModulators of cofactor recruitment by nuclear receptors are expected to play an important role in the coordination of hormone-induced transactivation processes. To identify such factors interacting with the N-terminal domain (NTD) of the progesterone receptor (PR), we used this domain as

  20. Niflumic acid-induced increase in potassium currents in frog motor nerve terminals: effects on transmitter release.

    Science.gov (United States)

    Miralles, F; Marsal, J; Peres, J; Solsona, C

    1996-04-01

    The actions of the nonsteroidal antiinflammatory drug niflumic acid were studied on frog neuromuscular preparations by conventional electrophysiological techniques. Niflumic acid reduced the amplitude and increased the latency of endplate potentials in a concentration-dependent manner. Neuromuscular junctions pretreated with niflumic acid (0.05-0.5 mM) showed much less depression than control when they were stimulated with trains of impulses. Inhibition of acetylcholine release was reverted by raising the extracellular Ca(2+) concentration but not by simply washing out the preparations with niflumic acid-free solutions. Pretreatment with indomethacin (0.1 mM), another nonsteroidal antiinflammatory drug, did not affect the niflumic acid-induced inhibition of evoked responses. Niflumic acid (0.1 mM) did not change the amplitude of miniature endplate potentials and had a dual action on the frequency of miniatures: it decreased their frequency at 0.1 mM whereas it produced an enormous increase in the rate of spontaneous discharge at 0.5 mM. Niflumic acid (0.1 - 1 mM) reversibly increased the amplitude and affected the kinetics of presynaptic voltage-activated K+ current and Ca(2+)-activated K(+) current in a concentration-dependent manner. Niflumic acid (0.1 - 1 mM) irreversibly decreased the amplitude and reversibly affected the kinetics of the nodal Na(+) current. Indomethacin (0.1 mM) had no effect on presynaptic currents. In conclusion, niflumic acid reduces acetylcholine release by increasing presynaptic K+ currents. This may shorten the depolarizing phase of the presynaptic action potential and may reduce the entry of Ca(2+) with each impulse.

  1. A20 zinc finger protein inhibits TNF-induced apoptosis and stress response early in the signaling cascades and independently of binding to TRAF2 or 14-3-3 proteins.

    Science.gov (United States)

    Lademann, U; Kallunki, T; Jäättelä, M

    2001-03-01

    A20 zinc finger protein is a negative regulator of tumor necrosis factor (TNF)-induced signaling pathways leading to apoptosis, stress response and inflammation. A20 has been shown to bind to TNF-receptor-associated factor 2 (TRAF2) and 14-3-3 chaperone proteins. Our data indicate that the zinc finger domain of A20 is sufficient and that neither TRAF2 nor 14-3-3 binding is necessary for the inhibitory effects of A20. Mutations in the 14-3-3 binding site of A20 did, however, result in a partial cleavage of A20 protein suggesting that 14-3-3 chaperone proteins may stabilize A20. Furthermore, we show that A20 acts early in TNF-induced signaling cascades blocking both TNF-induced rapid activation of c-Jun N-terminal kinase and processing of the receptor-associated caspase-8. Taken together our data indicate that the zinc finger domain of A20 contains all necessary functional domains required for the inhibition of TNF signaling and that A20 may function at the level of the receptor signaling complex.

  2. The terminal basal mitosis of chicken retinal Lim1 horizontal cells is not sensitive to cisplatin-induced cell cycle arrest.

    Science.gov (United States)

    Shirazi Fard, Shahrzad; Thyselius, Malin; All-Ericsson, Charlotta; Hallböök, Finn

    2014-01-01

    For proper development, cells need to coordinate proliferation and cell cycle-exit. This is mediated by a cascade of proteins making sure that each phase of the cell cycle is controlled before the initiation of the next. Retinal progenitor cells divide during the process of interkinetic nuclear migration, where they undergo S-phase on the basal side, followed by mitoses on the apical side of the neuroepithelium. The final cell cycle of chicken retinal horizontal cells (HCs) is an exception to this general cell cycle behavior. Lim1 expressing (+) horizontal progenitor cells (HPCs) have a heterogenic final cell cycle, with some cells undergoing a terminal mitosis on the basal side of the retina. The results in this study show that this terminal basal mitosis of Lim1+ HPCs is not dependent on Chk1/2 for its regulation compared to retinal cells undergoing interkinetic nuclear migration. Neither activating nor blocking Chk1 had an effect on the basal mitosis of Lim1+ HPCs. Furthermore, the Lim1+ HPCs were not sensitive to cisplatin-induced DNA damage and were able to continue into mitosis in the presence of γ-H2AX without activation of caspase-3. However, Nutlin3a-induced expression of p21 did reduce the mitoses, suggesting the presence of a functional p53/p21 response in HPCs. In contrast, the apical mitoses were blocked upon activation of either Chk1/2 or p21, indicating the importance of these proteins during the process of interkinetic nuclear migration. Inhibiting Cdk1 blocked M-phase transition both for apical and basal mitoses. This confirmed that the cyclin B1-Cdk1 complex was active and functional during the basal mitosis of Lim1+ HPCs. The regulation of the final cell cycle of Lim1+ HPCs is of particular interest since it has been shown that the HCs are able to sustain persistent DNA damage, remain in the cell cycle for an extended period of time and, consequently, survive for months.

  3. An agonistic monoclonal antibody against DR5 induces ROS production, sustained JNK activation and Endo G release in Jurkat leukemia cells

    Institute of Scientific and Technical Information of China (English)

    Caifeng Chen; Yanxin Liu; Dexian Zheng

    2009-01-01

    We have previously reported that AD5-10, a novel agonistic monoclonal antibody against DRS, possessed a strong cytotoxic activity in various tumor cells, via induction of caspase-dependent and-independent signaling pathways. The present study further demonstrates that reactive oxygen species (ROS) were generated in abundance in Jurkat leukemia cells upon ADS-10 stimulation and that ROS accumulation subsequently evoked sustained activation of c-Jun N-terminal kinase (JNK), loss of mitochondrial membrane potential, and release of endonuclease G (Endo G) from mitochondria into the cytosol. The reducing agent, N-acetylcysteine (NAC), effectively inhibited the sustained activation of JNK, release of Endo G, and cell death in Jurkat cells treated by ADS-10. Moreover, a dominant-nega-tive form of JNK (but not of p38) enhanced NF-KB activation, suppressed caspase-8 recruitment in death-inducing signaling complexes (DISCs), and reduced adverse effects on mitochondria, thereby inhibiting AD5-10-induced cell death in Jurkat leukemia cells. These data provide novel information on the DRS-mediated cell death-signaling path-way and may shed new light on effective strategies for leukemia and solid tumor therapies.

  4. The influence of acutely administered nicotine on cue-induced craving for gambling in at-risk video lottery terminal gamblers who smoke.

    Science.gov (United States)

    McGrath, Daniel S; Dorbeck, Anders; Barrett, Sean P

    2013-04-01

    Evidence indicates that tobacco use and gambling often co-occur. Despite this association, little is known about how tobacco use affects the propensity to gamble. Nicotine, the putative addictive component of tobacco, has been reported to potentiate the hedonic value of other nonsmoking stimuli. Environmental cues have been identified as an important contributor to relapse in addictive behavior; however, the extent to which nicotine can affect the strength of gambling cues remains unknown. This study examined whether nicotine influences subjective ratings for gambling following gambling cues. In a mixed within/between-subjects design, 30 (20 men) video lottery terminal (VLT) gamblers ('moderate-risk' or 'problem' gamblers) who smoke daily were assigned to nicotine (4 mg deliverable) or placebo lozenge conditions. Subjective and behavioral responses were assessed at baseline, following lozenge, following neutral cues, and following presentation of gambling cues. Nicotine lozenge was found to significantly reduce tobacco-related cravings (Pgambling-related cravings, the choice to play a VLT, or other subjective responses. These results suggest that a low dose of acutely administered nicotine does not increase cue-induced craving for gambling in at-risk VLT gamblers who smoke.

  5. Long term, continuous exposure to panobinostat induces terminal differentiation and long term survival in the TH-MYCN neuroblastoma mouse model.

    Science.gov (United States)

    Waldeck, Kelly; Cullinane, Carleen; Ardley, Kerry; Shortt, Jake; Martin, Ben; Tothill, Richard W; Li, Jason; Johnstone, Ricky W; McArthur, Grant A; Hicks, Rodney J; Wood, Paul J

    2016-07-01

    Neuroblastoma is the most common extra-cranial malignancy in childhood and accounts for ∼15% of childhood cancer deaths. Amplification of MYCN in neuroblastoma is associated with aggressive disease and predicts for poor prognosis. Novel therapeutic approaches are therefore essential to improving patient outcomes in this setting. The histone deacetylases are known to interact with N-Myc and regulate numerous cellular processes via epigenetic modulation, including differentiation. In this study, we used the TH-MYCN mouse model of neuroblastoma to investigate the antitumor activity of the pan-HDAC inhibitor, panobinostat. In particular we sought to explore the impact of long term, continuous panobinostat exposure on the epigenetically driven differentiation process. Continuous treatment of tumor bearing TH-MYCN transgenic mice with panobinostat for nine weeks led to a significant improvement in survival as compared with mice treated with panobinostat for a three-week period. Panobinostat induced rapid tumor regression with no regrowth observed following a nine-week treatment period. Initial tumor response was associated with apoptosis mediated via upregulation of BMF and BIM. The process of terminal differentiation of neuroblastoma into benign ganglioneuroma, with a characteristic increase in S100 expression and reduction of N-Myc expression, occurred following prolonged exposure to the drug. RNA-sequencing analysis of tumors from treated animals confirmed significant upregulation of gene pathways associated with apoptosis and differentiation. Together our data demonstrate the potential of panobinostat as a novel therapeutic strategy for high-risk neuroblastoma patients.

  6. Synthesis and application of a N-1' fluorescent biotinyl derivative inducing the specific carboxy-terminal dual labeling of a novel RhoB-selective scFv.

    Science.gov (United States)

    Chaisemartin, L; Chinestra, P; Favre, G; Blonski, C; Faye, J C

    2009-05-20

    The fluorescent site-specific labeling of protein would provide a new, easy-to-use alternative to biochemical and immunochemical methods. We used an intein-mediated strategy for covalent labeling of the carboxy-terminal amino acid of a RhoB-selective scFv previously isolated from a phage display library (a human synthetic V(H) + V(L) scFv phage library). The scFv fused to the Mxe intein was produced in E. coli and purified and was then labeled with a newly synthesized fluorescent biotinyl cysteine derivative capable of inducing scFv-Mxe intein splicing. In this study, we investigated the splicing and labeling properties of various amino acids in the hinge domain between scFv and Mxe under thiol activation. In this dual labeling system, the fluorescein is used for antibody detection and biotin is used for purification, resulting in a high specific activity for fluorescence. We then checked that the purified biotinylated fluorescent scFv retained its selectivity for RhoB without modification of its affinity.

  7. The archipelago ubiquitin ligase subunit acts in target tissue to restrict tracheal terminal cell branching and hypoxic-induced gene expression.

    Directory of Open Access Journals (Sweden)

    Nathan T Mortimer

    Full Text Available The Drosophila melanogaster gene archipelago (ago encodes the F-box/WD-repeat protein substrate specificity factor for an SCF (Skp/Cullin/F-box-type polyubiquitin ligase that inhibits tumor-like growth by targeting proteins for degradation by the proteasome. The Ago protein is expressed widely in the fly embryo and larva and promotes degradation of pro-proliferative proteins in mitotically active cells. However the requirement for Ago in post-mitotic developmental processes remains largely unexplored. Here we show that Ago is an antagonist of the physiologic response to low oxygen (hypoxia. Reducing Ago activity in larval muscle cells elicits enhanced branching of nearby tracheal terminal cells in normoxia. This tracheogenic phenotype shows a genetic dependence on sima, which encodes the HIF-1α subunit of the hypoxia-inducible transcription factor dHIF and its target the FGF ligand branchless (bnl, and is enhanced by depletion of the Drosophila Von Hippel Lindau (dVHL factor, which is a subunit of an oxygen-dependent ubiquitin ligase that degrades Sima/HIF-1α protein in metazoan cells. Genetic reduction of ago results in constitutive expression of some hypoxia-inducible genes in normoxia, increases the sensitivity of others to mild hypoxic stimulus, and enhances the ability of adult flies to recover from hypoxic stupor. As a molecular correlate to these genetic data, we find that Ago physically associates with Sima and restricts Sima levels in vivo. Collectively, these findings identify Ago as a required element of a circuit that suppresses the tracheogenic activity of larval muscle cells by antagonizing the Sima-mediated transcriptional response to hypoxia.

  8. C-terminal heat shock protein 90 inhibitor decreases hyperglycemia-induced oxidative stress and improves mitochondrial bioenergetics in sensory neurons.

    Science.gov (United States)

    Zhang, Liang; Zhao, Huiping; Blagg, Brian S J; Dobrowsky, Rick T

    2012-04-06

    Diabetic peripheral neuropathy (DPN) is a common complication of diabetes in which hyperglycemia-induced mitochondrial dysfunction and enhanced oxidative stress contribute to sensory neuron pathology. KU-32 is a novobiocin-based, C-terminal inhibitor of the molecular chaperone, heat shock protein 90 (Hsp90). KU-32 ameliorates multiple sensory deficits associated with the progression of DPN and protects unmyelinated sensory neurons from glucose-induced toxicity. Mechanistically, KU-32 increased the expression of Hsp70, and this protein was critical for drug efficacy in reversing DPN. However, it remained unclear if KU-32 had a broader effect on chaperone induction and if its efficacy was linked to improving mitochondrial dysfunction. Using cultures of hyperglycemically stressed primary sensory neurons, the present study investigated whether KU-32 had an effect on the translational induction of other chaperones and improved mitochondrial oxidative stress and bioenergetics. A variation of stable isotope labeling with amino acids in cell culture called pulse SILAC (pSILAC) was used to unbiasedly assess changes in protein translation. Hyperglycemia decreased the translation of numerous mitochondrial proteins that affect superoxide levels and respiratory activity. Importantly, this correlated with a decrease in mitochondrial oxygen consumption and an increase in superoxide levels. KU-32 increased the translation of Mn superoxide dismutase and several cytosolic and mitochondrial chaperones. Consistent with these changes, KU-32 decreased mitochondrial superoxide levels and significantly enhanced respiratory activity. These data indicate that efficacy of modulating molecular chaperones in DPN may be due in part to improved neuronal mitochondrial bioenergetics and decreased oxidative stress.

  9. A C-Terminal Heat Shock Protein 90 Inhibitor Decreases Hyperglycemia-induced Oxidative Stress and Improves Mitochondrial Bioenergetics in Sensory Neurons

    Science.gov (United States)

    Zhang, Liang; Zhao, Huiping; Blagg, Brian S.J.; Dobrowsky, Rick T.

    2012-01-01

    Summary Diabetic peripheral neuropathy (DPN) is a common complication of diabetes in which hyperglycemia-induced mitochondrial dysfunction and enhanced oxidative stress contribute to sensory neuron pathology. KU-32 is a novobiocin-based, C-terminal inhibitor of the molecular chaperone, heat shock protein 90 (Hsp90). KU-32 ameliorates multiple sensory deficits associated with the progression of DPN and protects unmyelinated sensory neurons from glucose-induced toxicity. Mechanistically, KU-32 increased the expression of Hsp70 and this protein was critical for drug efficacy in reversing DPN. However, it remained unclear if KU-32 had a broader effect on chaperone induction and if its efficacy was linked to improving mitochondrial dysfunction. Using cultures of hyperglycemically stressed primary sensory neurons, the present study investigated whether KU-32 had an effect on the translational induction of other chaperones and improved mitochondrial oxidative stress and bioenergetics. A variation of stable isotope labeling with amino acids in cell culture called pulse SILAC (pSILAC) was used to unbiasedly assess changes in protein translation. Hyperglycemia decreased the translation of numerous mitochondrial proteins that affect superoxide levels and respiratory activity. Importantly, this correlated with a decrease in mitochondrial oxygen consumption and an increase in superoxide levels. KU-32 increased the translation of Mn superoxide dismutase and several cytosolic and mitochondrial chaperones. Consistent with these changes, KU-32 decreased mitochondrial superoxide levels and significantly enhanced respiratory activity. These data indicate that efficacy of modulating molecular chaperones in DPN may be due in part to improved neuronal mitochondrial bioenergetics and decreased oxidative stress. PMID:22413817

  10. ERK 5/MAPK PATHWAY HAS A MAJOR ROLE IN 1α,25-(OH)2 VITAMIN D3-INDUCED TERMINAL DIFFERENTIATION OF MYELOID LEUKEMIA CELLS

    Science.gov (United States)

    Wang, Xuening; Pesakhov, Stella; Weng, Ashley; Kafka, Michael; Gocek, Elzbieta; Nguyen, Mai; Harrison, Jonathan S.; Danilenko, Michael; Studzinski, George P.

    2013-01-01

    Vitamin D derivatives, including its physiological form 1α,25(OH)2 vitamin D3 (1,25D), have anti-tumor actions demonstrated in cell culture and confirmatory epidemiological associations are frequently reported. However, their promise for use in the cancer clinic is still incompletely fulfilled, suggesting that a better understanding of the molecular events initiated by these compounds is needed for therapeutic advances. While ERK1/2 has been intensely investigated and is known to transmit signals for cell survival, growth, and differentiation, the role of other MAPK pathways has been studied sporadically. Therefore, we utilized acute myeloid leukemia (AML) cells in culture (HL60 and U937), to determine if ERK5 has a role in 1,25D-induced terminal differentiation which is distinct from the previously shown involvement of ERK1/2. We previously found that inhibition of kinase activity of ERK5 by specific pharmacological inhibitors BIX02189 or XMD8-92 results in higher expression of general myeloid marker CD11b, but a lower expression of the monocytic marker CD14. In contrast, the inhibition of the ERK1/2 pathway by PD98059 or U0126 reduced the expression of all differentiation markers studied. We report here for the first time that the differentiation changes induced by ERK5 inhibitors are accompanied by the inhibition of cell proliferation, and this occurs in the both G1 and G2 phases of the cell cycle. Of note, inhibition of ERK5 auto-phosphorylation by XMD8-92 results in a particularly robust cell cycle arrest in G2 phase in AML cells. This study provides a link between the 1,25D-elevated ERK5 pathway and changes in the cell cycle phase transitions in AML cells. Thus, combinations of vitamin D derivatives and ERK5 inhibitors may be more successful in cancer clinics than 1,25D or analogs alone. PMID:24514755

  11. Study on HepG-2 apoptosis induced by saponins isolated from Asparagus and the effects on the activities of caspase-3,8,9

    Institute of Scientific and Technical Information of China (English)

    JI Yu-bin; XU He; JI Chen-feng

    2008-01-01

    Objective To study the effect of saponins of asparagus on apoptosis and the variations of caspaseS, caspase-9 and caspase-3 activity in the process of asparagus induced apoptosis in HepG-2, to investigate the apoptosis mechanism further. Methods Asparagus on apoptosis effects on tumor cells cultured-HepG-2 with different concentrations at different time, IC50 value was measured by MTT assay, the apoptosis rate was determined by FCM with AnnexinV/PI staining, their apoptotic morphology were observed by electron microscopy and Colorimetric method was used to measure caspase-8,9 and caspase-3 activities. Results Experiments of antitumour in vivo showed that saponins of asparagus can inhibit the growth of tumor cell of HepG-2 in evidence, IC50 was 101.15 mg·L-1. Cultured for 72 h, the apoptosis rate had positive increased with concentrations. Apoptotic morphology was observed by electron microscopy. The activities of caspase-8, easpase-9 and caspase-3 had positive increased with concentrations. And have significant difference compared with negative control group(P<0.01). The activities of caspase-8 were high at 24 h, but the activities of caspase-9 and caspase-3 is high at 48 h. Conclusions Aaponins of asparagus can inhibit the growth of tumor cell of HepG2, and the underlying mechanism might be related to up regulation of caspase-8, 9 activity which subsequently transforms caspase-3 into its active form.

  12. Tyrosol prevents ischemia/reperfusion-induced cardiac injury in H9c2 cells: involvement of ROS, Hsp70, JNK and ERK, and apoptosis.

    Science.gov (United States)

    Sun, Liwei; Fan, Hang; Yang, Lingguang; Shi, Lingling; Liu, Yujun

    2015-02-25

    Ischemia-Reperfusion (I/R) injury causes ROS overproduction, creating oxidative stress, and can trigger myocyte death, resulting in heart failure. Tyrosol is an antioxidant abounded in diets and medicine. Our objective was to investigate the protective effect of tyrosol on I/R-caused mortality in H9c2 cardiomyocytes through its influence on ROS, Hsp70, ERK, JNK, Bcl-2, Bax and caspase-8. A simulated I/R model was used, myocytes loss was examined by MTT, and ROS levels were measured using DCFH-DA. Nuclear condensation and caspase-3 activity were assessed by DAPI staining and fluorometric assay. Phosphorylated ERK and JNK were determined by electrochemiluminescent ELISA, and Hsp70, Bcl-2, Bax and caspase-8 were examined by Western blotting. Results show that tyrosol salvaged myocyte loss, inhibited nuclear condensation and caspase-3 activity dose-dependently, indicating its protection against I/R-caused myocyte loss. Furthermore, tyrosol significantly inhibited ROS accumulation and activation of ERK and JNK, augmenting Hsp70 expression. Besides, tyrosol inhibited I/R-induced apoptosis, associated with retained anti-apoptotic Bcl-2 protein, and attenuated pro-apoptotic Bax protein, resulting in a preservation of Bcl-2/Bax ratio. Finally, tyrosol notably decreased cleaved caspase-8 levels. In conclusion, cytoprotection of tyrosol in I/R-caused myocyte mortality was involved with the mitigation of ROS, prohibition of the activation of ERK, JNK and caspase-8, and elevation of Hsp70 and Bcl-2/Bax ratio.

  13. Tyrosol Prevents Ischemia/Reperfusion-Induced Cardiac Injury in H9c2 Cells: Involvement of ROS, Hsp70, JNK and ERK, and Apoptosis

    Directory of Open Access Journals (Sweden)

    Liwei Sun

    2015-02-01

    Full Text Available Ischemia-Reperfusion (I/R injury causes ROS overproduction, creating oxidative stress, and can trigger myocyte death, resulting in heart failure. Tyrosol is an antioxidant abounded in diets and medicine. Our objective was to investigate the protective effect of tyrosol on I/R-caused mortality in H9c2 cardiomyocytes through its influence on ROS, Hsp70, ERK, JNK, Bcl-2, Bax and caspase-8. A simulated I/R model was used, myocytes loss was examined by MTT, and ROS levels were measured using DCFH-DA. Nuclear condensation and caspase-3 activity were assessed by DAPI staining and fluorometric assay. Phosphorylated ERK and JNK were determined by electrochemiluminescent ELISA, and Hsp70, Bcl-2, Bax and caspase-8 were examined by Western blotting. Results show that tyrosol salvaged myocyte loss, inhibited nuclear condensation and caspase-3 activity dose-dependently, indicating its protection against I/R-caused myocyte loss. Furthermore, tyrosol significantly inhibited ROS accumulation and activation of ERK and JNK, augmenting Hsp70 expression. Besides, tyrosol inhibited I/R-induced apoptosis, associated with retained anti-apoptotic Bcl-2 protein, and attenuated pro-apoptotic Bax protein, resulting in a preservation of Bcl-2/Bax ratio. Finally, tyrosol notably decreased cleaved caspase-8 levels. In conclusion, cytoprotection of tyrosol in I/R-caused myocyte mortality was involved with the mitigation of ROS, prohibition of the activation of ERK, JNK and caspase-8, and elevation of Hsp70 and Bcl-2/Bax ratio.

  14. Protocatechuic aldehyde inhibits TNF-α-induced fibronectin expression in human umbilical vein endothelial cells via a c-Jun N-terminal kinase dependent pathway.

    Science.gov (United States)

    Tong, Yue-Feng; Liu, Yong; Hu, Zhi-Xing; Li, Zhe-Cheng; A, Agula

    2016-01-01

    Fibronectin (FN) is one of the most important extracellular matrix proteins and plays an important role in the pathogenesis of atherosclerosis (AS). The aim of the present study was to evaluate the effect of a potent, water-soluble antioxidant, protocatechuic aldehyde (PA), which is derived from the Chinese herb Salvia miltiorrhiza, on the expression of FN in human umbilical vein endothelial cells (HUVECs) stimulated with tumor necrosis factor-α (TNF-α). The pharmacological effects of PA on the production of FN were investigated using ELISA and western blot analysis. In addition, ELISA and western blot analysis were used to examine the activation and suppression of the mitogen-activated protein kinase (MAPK) pathways and nuclear factor (NF)-κB in TNF-α-stimulated HUVECs, in order to explore the underlying pharmacological mechanism of PA. The inhibitory effect of PA on the total generation of reactive oxygen species (ROS) in TNF-α-stimulated HUVECs was assessed using 2',7'-dichlorofluorescein diacetate. Pretreatment of HUVECs with PA (0.15, 0.45 and 1.35 mM) for 18 h markedly attenuated the TNF-α-stimulated FN surface expression and secretion in a dose-dependent manner. Intracellular ROS generation and the expression of extracellular signal-regulated kinase 1 and 2 (ERK1/2), c-Jun N-terminal kinase (JNK) and p38 MAPK (p38) were significantly induced by TNF-α (2 ng/ml) in HUVECs. TNF-α-induced ROS generation and JNK activation were inhibited by PA in a concentration-dependent manner. By contrast, ERK1/2 and p38 activation was not significantly affected by PA. Pretreatment of HUVECs with PA for 18 h markedly attenuated TNF-α-stimulated NF-κB activation. In conclusion, the present findings suggest that PA inhibits TNF-α-induced FN expression in HUVECs through a mechanism that involves ROS/JNK and NF-κB.

  15. Docosahexaenoic acid sensitizes colon cancer cells to sulindac sulfide-induced apoptosis.

    Science.gov (United States)

    Lim, Soo-Jeong; Lee, Eunmyong; Lee, Eun-Hye; Kim, Soo-Yeon; Cha, Jun Hyung; Choi, Hwanho; Park, Wanseo; Choi, Hyeon Kyeom; Ko, Seong-Hee; Kim, So Hee

    2012-06-01

    Sulindac analogs represent one of the most efficacious groups of NSAIDs reducing the risk of colon cancer. Recent studies have shown that sulindac sulfide, a sulindac analog effective at lower doses compared to its parent compound, triggers the death receptor (DR)5-dependent extrinsic apoptotic pathway. Induction of apoptosis via activation of the DR-mediated pathway would be an ideal therapeutic strategy to eliminate cancer cells. In this study, we investigated the possibility that colon cancer cells are sensitized to sulindac sulfide-induced apoptosis by docosahexaenoic acid (DHA), via activation of the DR/extrinsic apoptotic pathway. Our data demonstrated that DHA combination sensitized colon cancer cells to sulindac sulfide-induced apoptosis, leading to enhanced growth suppression of human colon cancer xenografts. The combination effect was primarily attributed to increased cleavage of poly(ADP-ribose) polymerase (PARP) and caspase-8 activation. Moreover, pretreatment with z-IETD-FMK (caspase-8 inhibitor) or stable expression of dominant negative caspase-8 genes blocked DHA/sulindac sulfide cotreatment-induced apoptosis. In view of the finding that DR5 silencing abrogated the combination-stimulated apoptosis, we propose that apoptotic synergy induced by sulindac sulfide plus DHA is mediated via DR5. Our findings collectively support the utility of a combination of sulindac sulfide and DHA in the effective prevention and treatment of colon cancer.

  16. 2'-Nitroflavone induces apoptosis and modulates mitogen-activated protein kinase pathways in human leukaemia cells.

    Science.gov (United States)

    Cárdenas, Mariano G; Blank, Viviana C; Marder, Mariel N; Roguin, Leonor P

    2012-09-01

    The cytotoxic activity of 2'-nitroflavone was evaluated in different haematological cancer cell lines and its mechanism of action was further studied in HL-60 cells. 2'-Nitroflavone arrested the cell cycle at the G(2)/M phase and induced an apoptotic response characterized by an increase in the sub-G1 fraction of cells, a typical DNA ladder fragmentation, chromatin condensation and the detection of cells stained with Annexin V. Apoptosis was dependent on the activation of at least caspase-8, caspase-9 and caspase-3. The involvement of the death receptor pathway was indicated by the upregulation of both the tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) and its death receptor (DR5). We also showed that 2'-nitroflavone increased the expression levels of Bax and induced the release of cytochrome C to cytosol, suggesting the participation of the mitochondria-dependent pathway. When mitogen-activated protein kinases pathways were studied, it was found that p38 and c-Jun NH(2)-terminal kinase (JNK) pathways were activated by 2'-nitroflavone in HL-60 cells, whereas the phosphorylation levels of extracellular signal-regulated kinases (ERK) 1/2 decreased significantly. In addition, whereas both pharmacological inhibition of JNK and downregulation of JNK expression by RNA interference reduced the nitroflavone growth-inhibitory activity and the apoptotic effect, contrasting results were obtained when the ERK1/2 pathway was inhibited, and no effect was observed in the presence of a specific inhibitor of p38 mitogen-activated protein kinase. These findings show for the first time the antitumour action of 2'-nitroflavone in haematological cancer cell lines and suggest that both JNK and ERK1/2 cascades are involved in the apoptotic response induced by 2'-nitroflavone in HL-60 cells.

  17. Aging, Terminal Decline, and Terminal Drop

    Science.gov (United States)

    Palmore, Erdman; Cleveland, William

    1976-01-01

    Data from a 20-year longitudinal study of persons over 60 were analyzed by step-wise multiple regression to test for declines in function with age, for terminal decline (linear relationship to time before death), and for terminal drop (curvilinear relationship to time before death). There were no substantial terminal drop effects. (Author)

  18. Mechanisms Underlying Apoptosis-Inducing Effects of Kaempferol in HT-29 Human Colon Cancer Cells

    OpenAIRE

    Hyun Sook Lee; Han Jin Cho; Rina Yu; Ki Won Lee; Hyang Sook Chun; Jung Han Yoon Park

    2014-01-01

    We previously noted that kaempferol, a flavonol present in vegetables and fruits, reduced cell cycle progression of HT-29 cells. To examine whether kaempferol induces apoptosis of HT-29 cells and to explore the underlying molecular mechanisms, cells were treated with various concentrations (0–60 μmol/L) of kaempferol and analyzed by Hoechst staining, Annexin V staining, JC-1 labeling of the mitochondria, immunoprecipitation, in vitro kinase assays, Western blot analyses, and caspase-8 assays...

  19. Canine distemper virus induces apoptosis in cervical tumor derived cell lines

    Directory of Open Access Journals (Sweden)

    Rajão Daniela S

    2011-06-01

    Full Text Available Abstract Apoptosis can be induced or inhibited by viral proteins, it can form part of the host defense against virus infection, or it can be a mechanism for viral spread to neighboring cells. Canine distemper virus (CDV induces apoptotic cells in lymphoid tissues and in the cerebellum of dogs naturally infected. CDV also produces a cytopathologic effect, leading to apoptosis in Vero cells in tissue culture. We tested canine distemper virus, a member of the Paramyxoviridae family, for the ability to trigger apoptosis in HeLa cells, derived from cervical cancer cells resistant to apoptosis. To study the effect of CDV infection in HeLa cells, we examined apoptotic markers 24 h post infection (pi, by flow cytometry assay for DNA fragmentation, real-time PCR assay for caspase-3 and caspase-8 mRNA expression, and by caspase-3 and -8 immunocytochemistry. Flow cytometry showed that DNA fragmentation was induced in HeLa cells infected by CDV, and immunocytochemistry revealed a significant increase in the levels of the cleaved active form of caspase-3 protein, but did not show any difference in expression of caspase-8, indicating an intrinsic apoptotic pathway. Confirming this observation, expression of caspase-3 mRNA was higher in CDV infected HeLa cells than control cells; however, there was no statistically significant change in caspase-8 mRNA expression profile. Our data suggest that canine distemper virus induced apoptosis in HeLa cells, triggering apoptosis by the intrinsic pathway, with no participation of the initiator caspase -8 from the extrinsic pathway. In conclusion, the cellular stress caused by CDV infection of HeLa cells, leading to apoptosis, can be used as a tool in future research for cervical cancer treatment and control.

  20. Myocardial infarction-induced N-terminal fragment of cardiac myosin-binding protein C (cMyBP-C) impairs myofilament function in human myocardium.

    Science.gov (United States)

    Witayavanitkul, Namthip; Ait Mou, Younss; Kuster, Diederik W D; Khairallah, Ramzi J; Sarkey, Jason; Govindan, Suresh; Chen, Xin; Ge, Ying; Rajan, Sudarsan; Wieczorek, David F; Irving, Thomas; Westfall, Margaret V; de Tombe, Pieter P; Sadayappan, Sakthivel

    2014-03-28

    Myocardial infarction (MI) is associated with depressed cardiac contractile function and progression to heart failure. Cardiac myosin-binding protein C, a cardiac-specific myofilament protein, is proteolyzed post-MI in humans, which results in an N-terminal fragment, C0-C1f. The presence of C0-C1f in cultured cardiomyocytes results in decreased Ca(2+) transients and cell shortening, abnormalities sufficient for the induction of heart failure in a mouse model. However, the underlying mechanisms remain unclear. Here, we investigate the association between C0-C1f and altered contractility in human cardiac myofilaments in vitro. To accomplish this, we generated recombinant human C0-C1f (hC0C1f) and incorporated it into permeabilized human left ventricular myocardium. Mechanical properties were studied at short (2 μm) and long (2.3 μm) sarcomere length (SL). Our data demonstrate that the presence of hC0C1f in the sarcomere had the greatest effect at short, but not long, SL, decreasing maximal force and myofilament Ca(2+) sensitivity. Moreover, hC0C1f led to increased cooperative activation, cross-bridge cycling kinetics, and tension cost, with greater effects at short SL. We further established that the effects of hC0C1f occur through direct interaction with actin and α-tropomyosin. Our data demonstrate that the presence of hC0C1f in the sarcomere is sufficient to induce depressed myofilament function and Ca(2+) sensitivity in otherwise healthy human donor myocardium. Decreased cardiac function post-MI may result, in part, from the ability of hC0C1f to bind actin and α-tropomyosin, suggesting that cleaved C0-C1f could act as a poison polypeptide and disrupt the interaction of native cardiac myosin-binding protein C with the thin filament.

  1. Mechanisms underlying apoptosis-inducing effects of Kaempferol in HT-29 human colon cancer cells.

    Science.gov (United States)

    Lee, Hyun Sook; Cho, Han Jin; Yu, Rina; Lee, Ki Won; Chun, Hyang Sook; Park, Jung Han Yoon

    2014-02-17

    We previously noted that kaempferol, a flavonol present in vegetables and fruits, reduced cell cycle progression of HT-29 cells. To examine whether kaempferol induces apoptosis of HT-29 cells and to explore the underlying molecular mechanisms, cells were treated with various concentrations (0-60 μmol/L) of kaempferol and analyzed by Hoechst staining, Annexin V staining, JC-1 labeling of the mitochondria, immunoprecipitation, in vitro kinase assays, Western blot analyses, and caspase-8 assays. Kaempferol increased chromatin condensation, DNA fragmentation and the number of early apoptotic cells in HT-29 cells in a dose-dependent manner. In addition, kaempferol increased the levels of cleaved caspase-9, caspase-3 and caspase-7 as well as those of cleaved poly (ADP-ribose) polymerase. Moreover, it increased mitochondrial membrane permeability and cytosolic cytochrome c concentrations. Further, kaempferol decreased the levels of Bcl-xL proteins, but increased those of Bik. It also induced a reduction in Akt activation and Akt activity and an increase in mitochondrial Bad. Additionally, kaempferol increased the levels of membrane-bound FAS ligand, decreased those of uncleaved caspase-8 and intact Bid and increased caspase-8 activity. These results indicate that kaempferol induces the apoptosis of HT-29 cells via events associated with the activation of cell surface death receptors and the mitochondrial pathway.

  2. Laminarin-induced apoptosis in human colon cancer LoVo cells.

    Science.gov (United States)

    Ji, Chen-Feng; Ji, Yu-Bin

    2014-05-01

    A number of scientific studies have revealed that laminarin has antitumor effects. Therefore, the aim of the present study was to investigate the apoptosis of LoVo cells and the underlying mechanisms induced by laminarin. LoVo cells were treated with various concentrations of laminarin and fluorescence-inverted microscopy was used to observe the morphology of LoVo cells treated with laminarin. In addition, western blotting was performed to analyze the expression levels of death receptor (DR)4, DR5, TNF-related apoptosis-inducing ligand (TRAIL), Fas-associated protein with death domain (FADD), caspase-8, caspase-3, Bid and tBid. Flow cytometry was conducted to analyze the expressions of Bcl-2 and Bax, and spectrophotometry was performed to quantify the activity of caspases-8, -3, -6 and -7. Following the treatment of LoVo cells with laminarin for 24 h, the expression levels of DR4, DR5, TRAIL, FADD, Bid, tBid and Bax were observed to be upregulated, whereas the expression levels of pro-caspase-8, pro-caspase-3 and Bcl-2 were downregulated. In addition, the activities of casapse-8, -3, -6 and -7 were observed to increase, which was a significant difference when compared with those of the control group. Therefore, laminarin is considered to induce the apoptosis of LoVo cells, which may occur via a DR pathway, suggesting that laminarin may be a potent agent for cancer treatment.

  3. Gomisin N enhances TRAIL-induced apoptosis via reactive oxygen species-mediated up-regulation of death receptors 4 and 5

    OpenAIRE

    INOUE, Hiroki; WAIWUT, PORNTHIP; Saiki, Ikuo; Shimada, Yutaka; Sakurai, Hiroaki

    2011-01-01

    Pharmacological studies have revealed that lignans isolated from Schisandra chinensis, including gomisin N, show anticancer, anti-hepatotoxic, anti-oxidative and anti-inflammatory activities. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is an important member of the tumor necrosis factor superfamily with great potential in cancer therapy. The present study investigated whether pretreatment with gomisin N significantly enhanced TRAIL-induced cleavage of caspase-3, caspase-8 ...

  4. A methyl jasmonate derivative, J-7, induces apoptosis in human hepatocarcinoma Hep3B cells in vitro.

    Science.gov (United States)

    Park, Cheol; Jin, Cheng-Yun; Kim, Gi-Young; Cheong, JaeHun; Jung, Jee H; Yoo, Young Hyun; Choi, Yung Hyun

    2010-10-01

    The pro-apoptotic activity of J-7, a synthetic methyl jasmonate derivative, on the Hep3B human hepatocarcinoma cell line was investigated. Treatment of Hep3B cells with J-7 resulted in growth inhibition and the induction of apoptosis as measured by trypan blue-excluding cells, MTT assay, nuclear staining, DNA fragmentation, and flow cytometry analysis. The increased apoptotic events in Hep3B cells caused by J-7 were associated with the alteration in the ratio of Bax/Bcl-2 protein expression. J-7 treatment induced the expression of death receptor-related proteins such as death receptor 5, which triggered the activation of caspase-8 and the down-regulation of the whole Bid expression. In addition, the apoptosis induction by J-7 was correlated with the activation of caspase-9 and caspase-3, down-regulation IAP family proteins such as XIAP and cIAP-1, and concomitant degradation of poly (ADP-ribose) polymerase. However, the cytotoxic and apoptotic effects induced by J-7 were significantly inhibited by z-DEVD-fmk, a caspase-3 inhibitor, which demonstrates the important role that caspase-3 plays in the process. Furthermore, blocking the extracellular signal-regulated protein kinase and c-Jun N-terminal kinase pathways showed increased apoptosis and the activation of caspases in J-7-induced apoptosis. The results indicated that J-7 induces the apoptosis of Hep3B cells through a signaling cascade of death-receptor-mediated extrinsic as well as mitochondria-mediated intrinsic caspase pathways, which are associated with the activation of the mitogen-activated protein kinases signal pathway.

  5. A disruption of ctpA encoding carboxy-terminal protease attenuates Burkholderia mallei and induces partial protection in CD1 mice.

    Science.gov (United States)

    Bandara, Aloka B; DeShazer, David; Inzana, Thomas J; Sriranganathan, Nammalwar; Schurig, Gerhardt G; Boyle, Stephen M

    2008-09-01

    Burkholderia mallei is the etiologic agent of glanders in solipeds (horses, mules and donkeys), and incidentally in carnivores and humans. Little is known about the molecular mechanisms of B. mallei pathogenesis. The putative carboxy-terminal processing protease (CtpA) of B. mallei is a member of a novel family of endoproteases involved in the maturation of proteins destined for the cell envelope. All species and isolates of Burkholderia carry a highly conserved copy of ctpA. We studied the involvement of CtpA on growth, cell morphology, persistence, and pathogenicity of B. mallei. A sucrose-resistant strain of B. mallei was constructed by deleting a major portion of the sacB gene of the wild type strain ATCC 23344 by gene replacement, and designated as strain 23344DeltasacB. A portion of the ctpA gene (encoding CtpA) of strain 23344DeltasacB was deleted by gene replacement to generate strain 23344DeltasacBDeltactpA. In contrast to the wild type ATCC 23344 or the sacB mutant 23344DeltasacB, the ctpA mutant 23344DeltasacBDeltactpA displayed altered cell morphologies with partially or fully disintegrated cell envelopes. Furthermore, relative to the wild type, the ctpA mutant displayed slower growth in vitro and less ability to survive in J774.2 murine macrophages. The expression of mRNA of adtA, the gene downstream of ctpA was similar among the three strains suggesting that disruption of ctpA did not induce any polar effects. As with the wild type or the sacB mutant, the ctpA mutant exhibited a dose-dependent lethality when inoculated intraperitoneally into CD1 mice. The CD1 mice inoculated with a non-lethal dose of the ctpA mutant produced specific serum immunoglobulins IgG1 and IgG2a and were partially protected against challenge with wild type B. mallei ATCC 23344. These findings suggest that CtpA regulates in vitro growth, cell morphology and intracellular survival of B. mallei, and a ctpA mutant protects CD1 mice against glanders.

  6. β-Elemene piperazine derivatives induce apoptosis in human leukemia cells through downregulation of c-FLIP and generation of ROS.

    Directory of Open Access Journals (Sweden)

    Zhiying Yu

    Full Text Available β-Elemene is an active component of the herb medicine Curcuma Wenyujin with reported antitumor activity. To improve its antitumor ability, five novel piperazine derivatives of β-elemene, 13-(3-methyl-1-piperazinyl-β-elemene (DX1, 13-(cis-3,5-dimethyl-1-piperazinyl-β-elemene (DX2, 13-(4-ethyl-1-piperazinyl-β-elemene (DX3, 13-(4-isopropyl-1-piperazinyl-β-elemene (DX4 and 13-piperazinyl-β-elemene (DX5, were synthesized. The antiproliferative and apoptotic effects of these derivatives were determined in human leukemia HL-60, NB4, K562 and HP100-1 cells. DX1, DX2 and DX5, which contain a secondary amino moiety, were more active in inhibiting cell growth and in inducing apoptosis than DX3 and DX4. The apoptosis induction ability of DX1 was associated with the generation of hydrogen peroxide (H(2O(2, a decrease of mitochondrial membrane potential (MMP, and the activation of caspase-8. Pretreatment with the antioxidants N-acetylcysteine and catalase completely blocked DX1-induced H(2O(2 production, but only partially its activation of caspase-8 and induction of apoptosis. HL-60 cells were more sensitive than its H(2O(2-resistant subclone HP100-1 cells to DX1-induced apoptosis. The activation of caspase-8 by these compounds was correlated with the decrease in the levels of cellular FLICE-inhibitory protein (c-FLIP. The proteasome inhibitor MG-132 augmented the decrease in c-FLIP levels and apoptosis induced by these derivatives. FADD- and caspase-8-deficient Jurkat subclones have a decreased response to DX1-induced apoptosis. Our data indicate that these novel β-elemene piperazine derivatives induce apoptosis through the decrease in c-FLIP levels and the production of H(2O(2 which leads to activation of both death receptor- and mitochondrial-mediated apoptotic pathways.

  7. Ultrastructural changes in isolated peptidergic nerve terminals induced by digitonin permeabilization and K+ stimulation in the sinus gland of the crab, Cardisoma carnifex.

    Science.gov (United States)

    Nordmann, J J; Weatherby, T M; Haylett, B A

    1986-01-01

    The preparation of isolated peptidergic nerve terminals from the sinus gland (a neurohemal organ) of the crab (Cardisoma carnifex) is described. In this species the nerve endings can have diameters up to 30 microns. They release neurosecretory material as judged by the decrease in the volumetric density of granules upon depolarization with potassium. Similar results were obtained after permeabilization of the nerve terminals with digitonin, but only in the presence of micromolar concentrations of calcium. This preparation should prove useful in correlating electrical events with other cellular processes involved in stimulus-secretion coupling.

  8. Spontaneous and Fas-induced apoptosis of low-grade MDS erythroid precursors involves the endoplasmic reticulum.

    Science.gov (United States)

    Gyan, E; Frisan, E; Beyne-Rauzy, O; Deschemin, J-C; Pierre-Eugene, C; Randriamampita, C; Dubart-Kupperschmitt, A; Garrido, C; Dreyfus, F; Mayeux, P; Lacombe, C; Solary, E; Fontenay, M

    2008-10-01

    Spontaneous apoptosis of bone marrow erythroid precursors accounts for the anemia that characterizes most low-grade myelodysplastic syndromes (MDS). We have shown that death of these precursors involved the Fas-dependent activation of caspase-8. To explore the pathway leading from caspase-8 activation to apoptosis, we transduced MDS bone marrow CD34(+) cells with a lentivirus encoding wild-type (WT) or endoplasmic reticulum (ER)-targeted Bcl-2 protein before inducing their erythroid differentiation. Both WT-Bcl-2 and ER-targeted Bcl-2 prevented spontaneous and Fas-dependent apoptosis in MDS erythroid precursors. ER-targeted Bcl-2 inhibited mitochondrial membrane depolarization and cytochrome c release in MDS erythroid precursors undergoing apoptosis, indicating a role for the ER in the death pathway, upstream of the mitochondria. MDS erythroid precursors demonstrated elevated ER Ca(2+) stores and these stores remained unaffected by ER-targeted Bcl-2. The ER-associated protein Bcl-2-associated protein (BAP) 31 was cleaved by caspase-8 in MDS erythroid precursors undergoing apoptosis. The protective effect of ER-targeted Bcl-2 toward spontaneous and Fas-induced apoptosis correlated with inhibition of BAP31 cleavage. A protective effect of erythropoietin against Fas-induced BAP31 cleavage and apoptosis was observed. We propose that apoptosis of MDS erythroid precursors involves the ER, downstream of Fas and upstream of the mitochondria, through the cleavage of the ER-associated BAP31 protein.

  9. E2F activates late-G1 events but cannot replace E1A in inducing S phase in terminally differentiated skeletal muscle cells

    DEFF Research Database (Denmark)

    Pajalunga, D; Tognozzi, D; Tiainen, M

    1999-01-01

    We have previously shown that the adenovirus E1A oncogene can reactivate the cell cycle in terminally differentiated cells. Current models imply that much or all of this E1A activity is mediated by the release of the E2F transcription factors from pocket-protein control. In contrast, we show here...

  10. Procollagen type I N-terminal propeptide (PINP) as an indicator of type I collagen metabolism: ELISA development, reference interval, and hypovitaminosis D induced hyperparathyroidism

    DEFF Research Database (Denmark)

    Orum, O; Hansen, M; Jensen, Charlotte Harken;

    1996-01-01

    A sandwich enzyme-linked immunosorbent assay (ELISA) for quantification of the N-terminal propeptide of human procollagen type I (PINP) utilizing purified alpha 1-chain specific rabbit antibodies is described. The ELISA measured the content of the alpha 1-chain of PINP independent of the molecula...

  11. Lobaplatin suppresses proliferation and induces apoptosis in the human colorectal carcinoma cell Line LOVO in vitro.

    Science.gov (United States)

    Dai, Hong-yu; Liu, Lin; Qin, Shu-kui; He, Xiang-ming; Li, Su-yi

    2011-06-01

    Lobaplatin, as the third-generation platinum antineoplastic agent, showed promising antineoplastic effects in variety of preclinical test tumor models. We investigated the inhibition effect of lobaplatin on the colorectal carcinoma cell line LOVO in vitro, and explored its mechanism of action. The MTT assay was used to determine the inhibitory effect and inhibition ratio of lobaplatin on LOVO at various lobaplatin concentrations (500 μM, 1000 μM, 2000 μM). Apoptosis was detected by terminal deoxynucleotide transferase-mediated dUTP nickend labelling (TUNEL). The cell cycle and apoptotic rate were analyzed by flow cytometry (FCM) and the expression of caspase-3,8,9 in cells was detected by chromometry. The results of MTT assay showed that proliferation of LOVO cells was inhibited by lobaplatin in a concentration-dependent manner. Apoptosis was detected in LOVO cells by TUNEL. The FCM assay indicated that lobaplatin altered the cell cycle and induced apoptosis of the LOVO cells when treated for 24h, the percentages of cells in the S phase transition were increased, whereas the percentages of cells in the G(2) transition were decreased. The expressions of caspase-389 is higher than the control group after LOVO cells were treated by lobaplatin. Lobaplatin can inhibit the proliferation of colorectal carcinoma cell line LOVO by inducing apoptosis in vitro. The mechanism may be related to the "S" cycle arrest in cell cycle distribution and the up-regulated expression of caspase-8 and caspase-9 which up-regulated the expression of caspase-3.

  12. Caspase cleavage product of BAP31 induces mitochondrial fission through endoplasmic reticulum calcium signals, enhancing cytochrome c release to the cytosol.

    Science.gov (United States)

    Breckenridge, David G; Stojanovic, Marina; Marcellus, Richard C; Shore, Gordon C

    2003-03-31

    Stimulation of cell surface death receptors activates caspase-8, which targets a limited number of substrates including BAP31, an integral membrane protein of the endoplasmic reticulum (ER). Recently, we reported that a caspase-resistant BAP31 mutant inhibited several features of Fas-induced apoptosis, including the release of cytochrome c (cyt.c) from mitochondria (Nguyen, M., D.G. Breckenridge, A. Ducret, and G.C. Shore. 2000. Mol. Cell. Biol. 20:6731-6740), implicating ER-mitochondria crosstalk in this pathway. Here, we report that the p20 caspase cleavage fragment of BAP31 can direct pro-apoptotic signals between the ER and mitochondria. Adenoviral expression of p20 caused an early release of Ca2+ from the ER, concomitant uptake of Ca2+ into mitochondria, and mitochondrial recruitment of Drp1, a dynamin-related protein that mediates scission of the outer mitochondrial membrane, resulting in dramatic fragmentation and fission of the mitochondrial network. Inhibition of Drp1 or ER-mitochondrial Ca2+ signaling prevented p20-induced fission of mitochondria. p20 strongly sensitized mitochondria to caspase-8-induced cyt.c release, whereas prolonged expression of p20 on its own ultimately induced caspase activation and apoptosis through the mitochondrial apoptosome stress pathway. Therefore, caspase-8 cleavage of BAP31 at the ER stimulates Ca2+-dependent mitochondrial fission, enhancing the release of cyt.c in response to this initiator caspase.

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    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  6. krwl Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  7. kpsp Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  8. kcmi Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  9. ktlh Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  10. kteb Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  11. kbis Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  12. kagc Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  13. krbg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  14. kelm Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  15. khln Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  16. kgeg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  17. kazo Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  18. klan Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  19. ksmn Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  20. klbx Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  1. kbvi Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  2. ksjc Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  3. klnd Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  4. klru Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  5. kbrl Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  6. pawg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  7. klaf Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  8. kgon Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  9. ksdy Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  10. ksea Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  11. kofk Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  12. krdg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  13. kdug Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  14. kbwg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  15. kbbg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  16. krdu Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  17. kiwd Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  18. krbl Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  19. kssf Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  20. ksaw Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  1. kmot Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  2. kiso Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  3. kgck Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  4. kcvg Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  5. pafa Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  6. kcrq Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  7. ksun Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  8. khdn Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  9. pabe Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  10. ktop Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  11. ktrk Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  12. krvs Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  13. phny Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  14. ktcl Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  15. kmrb Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  16. kcub Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  17. kpdt Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  18. ptkk Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  19. kcsm Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  20. ksfo Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  1. kcgi Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  2. kjhw Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  3. kdab Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  4. kecp Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  5. kmls Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  6. kroc Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  7. kfod Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  8. kmia Terminal Aerodrome Forecast

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — TAF (terminal aerodrome forecast or terminal area forecast) is a format for reporting weather forecast information, particularly as it relates to aviation. TAFs are...

  9. 49 CFR 1242.27 - Coal marine terminals, ore marine terminals, TOFC/COFC terminals, other marine terminals, motor...

    Science.gov (United States)

    2010-10-01

    ... 49 Transportation 9 2010-10-01 2010-10-01 false Coal marine terminals, ore marine terminals, TOFC/COFC terminals, other marine terminals, motor vehicle loading and distribution facilities, and... Structures § 1242.27 Coal marine terminals, ore marine terminals, TOFC/COFC terminals, other marine...

  10. Organizational Relationship Termination Competence

    DEFF Research Database (Denmark)

    Ritter, Thomas; Geersbro, Jens

    2011-01-01

    that a firm's percentage of unwanted customers decreases significantly as acceptance of termination increases, if the firm's definition of unwanted customers is well understood, and if a firm has clear termination routines. In addition, general focus on profitability and external constraints on relationship...... termination are found to significantly affect a firm's relationship termination competence. The findings suggest that managers should regard termination as a legitimate option in customer relationship management. In order to decrease the number of unwanted customers, managers must accept termination......Most firms are involved in a number of customer relationships that drain the firm's resources. However, many firms are hesitant to address this problem. This paper investigates customer relationship termination at the organizational level. We develop and analyze the organizational dimensions...

  11. CONTAINER TERMINALS IN EUROPE

    Directory of Open Access Journals (Sweden)

    Bart W. WIEGMANS

    2001-01-01

    Full Text Available This paper aims to address the linkage between logistics (in particular, the management of marketing channel flows and transport markets, while also the interaction between these two markets and intermodal container terminals is analysed. The marketing channel theory is used to describe all relevant actors and flows that run through marketing channels, starting with customer needs and ending with customer satisfaction. Porter's theory of competitive advantages is used to review competitive forces in both markets. Finally, a competitor analysis is performed for the logistics and transport market. These theories are applied so as to be able to determine the competitive position of intermodal container terminals with a view to the management of marketing channel flows and the physical transport of freight flows. Hence, the central question of this paper is: Which markets are served by intermodal container terminals and with whom are they competing? At present, neither the maritime container terminals nor the continental container terminals appear to have a significant influence in the logistics service market; they concentrate mainly on the physical movement of containers (transshipment. Furthermore, maritime container terminals and continental container terminals are not dominant players in the transport service market. Our conclusion is that continental terminals are predominantly competing with unimodal road transport, with neighbouring continental terminals and with barge transport companies.

  12. Inhibition of c-Jun N-terminal Kinase Signaling Pathway Alleviates Lipopolysaccharide-induced Acute Respiratory Distress Syndrome in Rats

    Directory of Open Access Journals (Sweden)

    Jian-Bo Lai

    2016-01-01

    Conclusions: Inhibiting JNK alleviated LPS-induced acute lung inflammation and had no effects on pulmonary edema and fibrosis. JNK inhibitor might be a potential therapeutic medication in ARDS, in the context of reducing lung inflammatory.

  13. Stress-induced phosphorylation of c-Jun-N-terminal kinases and nuclear translocation of Hsp70 in the Wistar rat hippocampus

    Directory of Open Access Journals (Sweden)

    Adžić M.

    2009-01-01

    Full Text Available Glucocorticoids are key regulators of the neuroendocrine stress response in the hippocampus. Their action is partly mediated through the subfamily of MAPKs termed c-Jun-N-terminal kinases (JNKs,whose activation correlates with neurodegeneration. The stress response also involves activation of cell protective mechanisms through various heat shock proteins (HSPs that mediate neuroprotection. We followed both JNKs and Hsp70 signals in the cytoplasmic and nuclear compartments of the hippocampus of Wistar male rats exposed to acute, chronic, and combined stress. The activity of JNK1 was decreased in both compartments by all three types of stress, while the activity of cytoplasmic JNK2/3 was elevated in acute and unaltered or lowered in chronic and combined stress. Under all stress conditions, Hsp70 translocation to the nucleus was markedly increased. The results suggest that neurodegenerative signaling of JNKs may be counteracted by increase of nuclear Hsp70,especially under chronic stress.

  14. Two millennia of tropical cyclone-induced mud layers in a northern Yucatán stalagmite: Multiple overlapping climatic hazards during the Maya Terminal Classic "megadroughts"

    Science.gov (United States)

    Frappier, Amy Benoit; Pyburn, James; Pinkey-Drobnis, Aurora D.; Wang, Xianfeng; Corbett, D. Reide; Dahlin, Bruce H.

    2014-07-01

    An annually laminated stalagmite from the northern Yucatán Peninsula contains mud layers from 256 cave flooding events over 2240 years. This new conservative proxy for paleotempestology recorded cave flooding events with a recurrence interval of 8.3 years during the twentieth century, with the greatest frequency during the twentieth century and the least frequent during the seventeenth century. Tropical cyclone (TC) events are unlikely to flood the cave during drought when the water table is depressed. Applying TC masking to the Chaac paleorainfall reconstruction suggests that the severity of the Maya "megadroughts" was underestimated. Without a high-resolution radiometric geochronology of individual local TC events, speleothem isotope records cannot resolve whether the Terminal Classic Period in the northern Maya Lowlands was punctuated by several brief drought breaks with normal TCs, or whether the region was very dry and peppered by unusually severe and frequent hurricane seasons.

  15. Effects of p53 on fisetin-induced apoptosis in hepatocellular carcinoma cell line HepG2%p53在漆树黄酮诱导人肝癌细胞系HepG2细胞凋亡的作用

    Institute of Scientific and Technical Information of China (English)

    李蓉; 黎小兵; 蔡康荣; 陈锦; 黄培春

    2011-01-01

    目的 观察漆树黄酮诱导人肝癌细胞系HepG2细胞凋亡的现象,研究p53在漆树黄酮诱导人肝癌细胞系HepG2细胞凋亡的作用.方法 用不同浓度的漆树黄酮(50、100、200 μmol/L)处理HepG2细胞24 h和100 μmol/L漆树黄酮处理HepG2细胞不同时间(12、24、48 h),流式细胞技术和Hoechst/PI免疫荧光法观察漆树黄酮作用HepG2细胞的凋亡情况; Westem Blot法检测漆树黄酮对HepG2细胞P53、Caspase-3 和Caspase-8蛋白的表达情况.结果 漆树黄酮能诱导HepG2细胞凋亡,呈剂量-时间依赖性; 漆树黄酮可增加P53蛋白的表达,降解Caspase-3、Caspase-8酶原,提高Caspase-3、Caspase-8活性.P53蛋白的上调与凋亡的发生呈正相关(r=0.586,r=0.592,均P<0.01),Caspase-3、Caspase-8酶原的降解与与凋亡的发生呈负相关(r=-0.585~-0.583,均P<0.01).结论 漆树黄酮可诱导HepG2细胞凋亡,其机制可能与上调p53表达,活化Caspase-3、 Caspase-8有关.%Objective To study the effects of p53 on fisetin - induced apoptosis in hepatocellular carcinoma cell line HepG2 cells. Methods HepG2 cells were treated with fisetin at different concentrations (50. 100, 200 μmol/L)for 24h and with fisetin of 100 μmol/L with different duration ( 12 , 24 , 48 h) . Cell apoptosis was assessed with Hoechst/PI double staining assay and flow cytometry ( FCM) . The expression levels of P53 , Caspase - 3 and Caspase - 8 proteins in HepG2 cells were measured by Western Blot. Results Fisetin - induced apoptosis in HepG2 cells followed dose - and time - dependent manners. Chromatin condensation was observed with Hoechst/PI staining in fisetin - treated cells. The level of P53 was significantly elevated in HepG2 cells after fisetin treatment, while the levels of Caspase - 3 and Caspase - 8 were significantly reduced. Significant positive correlation was observed between up - regulation of P53 and cellular apoptosis , while negative correlation was observed between down - regulation of

  16. Cadmium induces apoptosis in pancreatic β-cells through a mitochondria-dependent pathway: the role of oxidative stress-mediated c-Jun N-terminal kinase activation.

    Directory of Open Access Journals (Sweden)

    Kai-Chih Chang

    Full Text Available Cadmium (Cd, one of well-known highly toxic environmental and industrial pollutants, causes a number of adverse health effects and diseases in humans. The growing epidemiological studies have suggested a possible link between Cd exposure and diabetes mellitus (DM. However, the toxicological effects and underlying mechanisms of Cd-induced pancreatic β-cell injury are still unknown. In this study, we found that Cd significantly decreased cell viability, and increased sub-G1 hypodiploid cells and annexin V-Cy3 binding in pancreatic β-cell-derived RIN-m5F cells. Cd also increased intracellular reactive oxygen species (ROS generation and malondialdehyde (MDA production and induced mitochondrial dysfunction (the loss of mitochondrial membrane potential (MMP and the increase of cytosolic cytochrome c release, the decreased Bcl-2 expression, increased p53 expression, poly (ADP-ribose polymerase (PARP cleavage, and caspase cascades, which accompanied with intracellular Cd accumulation. Pretreatment with the antioxidant N-acetylcysteine (NAC effectively reversed these Cd-induced events. Furthermore, exposure to Cd induced the phosphorylations of c-jun N-terminal kinases (JNK, extracellular signal-regulated kinases (ERK1/2, and p38-mitogen-activated protein kinase (MAPK, which was prevented by NAC. Additionally, the specific JNK inhibitor SP600125 or JNK-specific small interference RNA (si-RNA transfection suppressed Cd-induced β-cell apoptosis and related signals, but not ERK1/2 and p38-MAPK inhibitors (PD98059 and SB203580 did not. However, the JNK inhibitor or JNK-specific si-RNA did not suppress ROS generation in Cd-treated cells. These results indicate that Cd induces pancreatic β-cell death via an oxidative stress downstream-mediated JNK activation-triggered mitochondria-regulated apoptotic pathway.

  17. A role for CD4 sup + but not CD8 sup + T cells in immunity to Schistosoma mansoni induced by 20 krad-irradiated and Ro 11-3128-terminated infections

    Energy Technology Data Exchange (ETDEWEB)

    Vignali, D.A.A.; Bickle, Q.D.; Taylor, M.G. (London School of Hygiene and Tropical Medicine (UK)); Crocker, P. (Oxford Univ. (UK). Sir William Dunn School of Pathology); Cobbold, S.; Waldmann, H (Cambridge Univ. (UK). Dept. of Pathology)

    1989-08-01

    The role of CD4{sup +} (L3/T4{sup +}) and CD8{sup +} (Lyt-2{sup +}) T cells in immunity to Schistosoma mansoni induced by 20 krad-irradiated and Ro 11-terminated infections in mice was investigated directly by in vivo depletion of these subsets with cytotoxic rat monoclonal antibodies (mAb). Effective physical depletion was demonstrated by flow cytometric analysis and immunohistochemical staining. Functional depletion of helper activity following anti-CD4 treatment was indicated by an abrogation of concanavalin A(Con A)-induced colony-stimulating factor (CSF) release, while anti-CD8 treatment had no effect in these assays. Pre-existing S. mansoni-specific antibody levels were unaffected by anti-CD4 and anti-CD8 treatment. In vivo depletion of CD4 {sup +} T cells resulted in a dramatic reduction in immunity induced by one (up to 100%) and two (up to 70%) vaccinations with 20 krad-irradiated cercariae and also of resistance induced by Ro 11-attenuated infections (up to 100%). Depletion of CD8{sup +} T cells had no effect on resistance induced by any of the vaccination protocols investigated. A correlation was observed between resistance and T cell-induced, macrophage-mediated killing of schistosomula in vitro, both of which were abrogated following anti-CD4 treatment but were unaffected by CD8{sup +} T-cell depletion. The possible role of CD4{sup +} T cells in vivo and the implications for vaccine development are discussed. (author).

  18. Growth arrest- and DNA-damage-inducible 45beta gene inhibits c-Jun N-terminal kinase and extracellular signal-regulated kinase and decreases IL-1beta-induced apoptosis in insulin-producing INS-1E cells

    DEFF Research Database (Denmark)

    Larsen, Claus Morten; Døssing, M G; Papa, S;

    2006-01-01

    IL-1beta is a candidate mediator of apoptotic beta cell destruction, a process that leads to type 1 diabetes and progression of type 2 diabetes. IL-1beta activates beta cell c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38, all of which are members of the mitogen...

  19. The Epstein-Barr virus (EBV) glycoprotein B cytoplasmic C-terminal tail domain regulates the energy requirement for EBV-induced membrane fusion.

    Science.gov (United States)

    Chen, Jia; Zhang, Xianming; Jardetzky, Theodore S; Longnecker, Richard

    2014-10-01

    The entry of enveloped viruses into host cells is preceded by membrane fusion, which in Epstein-Barr virus (EBV) is thought to be mediated by the refolding of glycoprotein B (gB) from a prefusion to a postfusion state. In our current studies, we characterized a gB C-terminal tail domain (CTD) mutant truncated at amino acid 843 (gB843). This truncation mutant is hyperfusogenic as monitored by syncytium formation and in a quantitative fusion assay and is dependent on gH/gL for fusion activity. gB843 can rescue the fusion function of other glycoprotein mutants that have null or decreased fusion activity in epithelial and B cells. In addition, gB843 requires less gp42 and gH/gL for fusion, and can function in fusion at a lower temperature than wild-type gB, indicating a lower energy requirement for fusion activation. Since a key step in fusion is the conversion of gB from a prefusion to an active postfusion state by gH/gL, gB843 may access this activated gB state more readily. Our studies indicate that the gB CTD may participate in the fusion function by maintaining gB in an inactive prefusion form prior to activation by receptor binding. Importance: Diseases resulting from Epstein-Barr virus (EBV) infection in humans range from the fairly benign disease infectious mononucleosis to life-threatening cancer. As an enveloped virus, EBV must fuse with a host cell membrane for entry and infection by using glycoproteins gH/gL, gB, and gp42. Among these glycoproteins, gB is thought to be the protein that executes fusion. To further characterize the function of the EBV gB cytoplasmic C-terminal tail domain (CTD) in fusion, we used a previously constructed CTD truncation mutant and studied its fusion activity in the context of other EBV glycoprotein mutants. From these studies, we find that the gB CTD regulates fusion by altering the energy requirements for the triggering of fusion mediated by gH/gL or gp42. Overall, our studies may lead to a better understanding of EBV fusion

  20. Contraction-induced Interleukin-6 Gene Transcription in Skeletal Muscle Is Regulated by c-Jun Terminal Kinase/Activator Protein-1*

    Science.gov (United States)

    Whitham, Martin; Chan, M. H. Stanley; Pal, Martin; Matthews, Vance B.; Prelovsek, Oja; Lunke, Sebastian; El-Osta, Assam; Broenneke, Hella; Alber, Jens; Brüning, Jens C.; Wunderlich, F. Thomas; Lancaster, Graeme I.; Febbraio, Mark A.

    2012-01-01

    Exercise increases the expression of the prototypical myokine IL-6, but the precise mechanism by which this occurs has yet to be identified. To mimic exercise conditions, C2C12 myotubes were mechanically stimulated via electrical pulse stimulation (EPS). We compared the responses of EPS with the pharmacological Ca2+ carrier calcimycin (A23187) because contraction induces marked increases in cytosolic Ca2+ levels or the classical IκB kinase/NFκB inflammatory response elicited by H2O2. We demonstrate that, unlike H2O2-stimulated increases in IL-6 mRNA, neither calcimycin- nor EPS-induced IL-6 mRNA expression is under the transcriptional control of NFκB. Rather, we show that EPS increased the phosphorylation of JNK and the reporter activity of the downstream transcription factor AP-1. Furthermore, JNK inhibition abolished the EPS-induced increase in IL-6 mRNA and protein expression. Finally, we observed an exercise-induced increase in both JNK phosphorylation and IL-6 mRNA expression in the skeletal muscles of mice after 30 min of treadmill running. Importantly, exercise did not increase IL-6 mRNA expression in skeletal muscle-specific JNK-deficient mice. These data identify a novel contraction-mediated transcriptional regulatory pathway for IL-6 in skeletal muscle. PMID:22351769

  1. Highly Oxygenated Sesquiterpene Lactones from Cousinia aitchisonii and their Cytotoxic Properties: Rhaserolide Induces Apoptosis in Human T Lymphocyte (Jurkat) Cells via the Activation of c-Jun n-terminal Kinase Phosphorylation.

    Science.gov (United States)

    Iranshahy, Milad; Tayarani-Najaran, Zahra; Kasaian, Jamal; Ghandadi, Morteza; Emami, Seyed Ahmad; Asili, Javad; Chandran, Jima N; Schneider, Bernd; Iranshahi, Mehrdad

    2016-02-01

    Infrared-guided chromatographic fractionation of sesquiterpene lactones from the extracts of Cousinia aitchisonii and Cousinia concolor led to the isolation of five pure compounds. A new sesquiterpene lactone, namely, aitchisonolide, and two known sesquiterpene lactones (desoxyjanerin and rhaserolide) were isolated from C. aitchisonii and two known lignans (arctiin and arctigenin) from C. concolor. The structures of these compounds were elucidated by one-dimensional and two-dimensional nuclear magnetic resonance techniques, as well as high-resolution mass spectrometry. The purified and characterized compounds were subjected to cytotoxicity assay. The sesquiterpene lactones desoxyjanerin and rhaserolide showed significant cytotoxic activities against five different cancer cell lines and the normal human embryonic kidney cell line. Rhaserolide was chosen to evaluate the possible mechanism of action. Western blot analysis revealed that rhaserolide could induce apoptosis in Jurkat cells via the activation of c-Jun n-terminal kinase phosphorylation.

  2. Abrupt reduction of c-myc expression by antisense RNA inducing terminal differentiation and apoptosis of a human esophageal cancer cell line

    Institute of Scientific and Technical Information of China (English)

    赵晓航; 王秀琴; 周传农; 彭仁玲; 阎水中; 吴旻

    1995-01-01

    A human esophageal cancer cell line (EC8712) expressing high-level Myc protein was infected with recombmant retroviral particles (pA-BD9) at a multiplicity of infection (MOI) 1:1. This viral particle contains a neomycin-resistant gene and a 1.53-kb antisense RNA spanning the 2nd exon and its flanking sequences of the human c-myc oncogene. The G418-resistant EC8712 clones showed an 86% inhibition of growth rate and morphological changes characteristic of terminal differentiation and apoptosis. A decrease of about 80% of Myc protein was also observed in these infected cells by ABC-ELISA assay. 12-24 h after the infection of ECS712 cells with pA-BD9 at a high viral particle concentration (MOI = 1:10), the integration of the extrinsic 1.53-kb antisense c-myc fragment into the cancer cell genome was evidenced by the Southern blot analysis. Northern blot analyses showed the expression of this antisense fragment and a decrease of the intrinsic c-myc expression by 74% in comparison with that of the parental EC8

  3. Effect of the human follicle-stimulating hormone-binding inhibitor and its N-terminal fragment on follicle-stimulating hormone-induced progesterone secretion by granulosa cells in vitro

    Indian Academy of Sciences (India)

    Perinaaz R Wadia; Smita D Mahale; Tarala D Nandedkar

    2007-09-01

    Intrafollicular factors play an important role in folliculogenesis. The follicle-stimulating hormone (FSH)-binding inhibitor (FSHBI), purified by our laboratory from human ovarian follicular fluid, has been shown to suppress ovulation and induce follicular atresia/apoptosis in mice as well as impair fertility in marmosets, the new world monkeys. The octapeptide, a peptide corresponding to the N-terminal region of human FSHBI (hFSHBI), has been synthesized and also shows FSHBI activity in vitro. In the present study, we have attempted to identify the mechanism of action of the peptide in granulosa cell cultures. Rat granulosa cell cultures were treated with varying concentrations of the octapeptide or partially purified hFSHBI (gel chromatography fraction hGF2) in the presence or absence of human FSH (hFSH) and the amount of progesterone (P4) secreted in the culture supernatants after 3 h/48 h was estimated. Both hGF2 and the octapeptide failed to alter basal levels as well as 8-bromo cAMP-induced P4 production, while FSH-induced P4 secretion was inhibited in a dose-dependent manner. These studies reveal that the octapeptide, a fragment of FSHBI, and the native protein have similar activity in vitro and both compounds alter FSH action at the receptor level upstream of cAMP formation.

  4. Downregulation of the Spi-1/PU.1 oncogene induces the expression of TRIM10/HERF1, a key factor required for terminal erythroid cell differentiation and survival

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Sustained expression of the Spi-1/PU.1 and Fli-1 oncoproteins blocks globin gene activation in mouse erythroleukemia cells; however, only Spi-1/PU.1 expression inhibits the inclusion of exon 16 in the mature 4.1R mRNA. This splicing event is crucial for a functional 4.1R protein and, therefore, for red blood cell membrane integrity. This report demonstrates that Spi-I/PU.1 downregulation induces the activation of TRiMl0/hematopoietic RING finger 1 (HERFI), a member of the tripartite motif (TRIM)/RBCC protein family needed for globin gene transcription. Additionally, we demonstrate that TRIM10/HERFI is required for the regulated splicing of exon 16 during late erythroid differentiation. Using inducible overexpression and silencing approaches, we found that: (1) TRIM10/HERF1 knockdown inhibits hemoglobin production and exon splicing and triggers cell apoptosis in dimethylsulfoxide (DMSO)-induced cells; (2) TRIMI0/HERF1 upregulation is required but is insufficient on its own to activate exon retention; (3) Fli-1 has no effect on TRIMI0/HERFI expression, whereas either DMSO-induced downregulation or shRNA-knockdown of Spi-1/PU.1 expression is sufficient to activate TRIM10/HERFI expression; and (4) Spi-1/PU.1 knockdown triggers both the transcription and the splicing events independently of the chemical induction. Altogether, these data indicate that primary Spi-1/PU.1 downregulation acts on late erythroid differentiation through at least two pathways, one of which requires TRIM10/HERF1 upregulation and parallels the Spi-1/PU.1-induced Fli-1 shutoff regulatory cascade.

  5. Metformin induces apoptosis of pancreatic cancer cells

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM: To assess the role and mechanism of mefformin in inducing apoptosis of pancreatic cancer cells. METHODS: The human pancreatic cancer cell lines ASPC-1, BxPc-3, PANC-1 and SW1990 were exposed to mefformin. The inhibition of cell proliferation and colony formation via apoptosis induction and S phase arrest in pancreatic cancer cell lines of mefformin was tested.RESULTS: In each pancreatic cancer cell line tested, metformin inhibited cell proliferation in a dose dependent manner in MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assays). Flow cytometric analysis showed that metformin reduced the number of cells in G1 and increased the percentage of cells in S phase as well as the apoptotic fraction. Enzymelinked immunosorbent assay (EUSA) showed that metformin induced apaptosis in all pancreatic cancer cell lines. In Western blot studies, metformin induced oly-ADP-ribose polymerase(PARP) cleavage (an indicator of aspase activation) in all pancreatic cancer cell lines. The general caspase inhibitor (VAD-fmk) completely abolished metformin-induced PARP cleavage and apoptosis in ASPC-1 BxPc-3 and PANC-1, the caspase-8 specific inhibitor (IETD-fmk) and the caspase-9 specific inhibitor (LEHD-fmk) only partially abrogated metformin-induced apoptosis and PARP cleavage in BxPc-3 and PANC-1 cells. We also observed that metformin treatment ramatically reduced epidermal growth factor receptor (EGFR) and phosphorylated mitogen activated protein kinase (P-MAPK) in both a time- and dose-dependent manner in all cell lines tested.CONCLUSION: Metformin significantly inhibits cell proliferation and apoptosis in all pancreatic cell lines. And the metformin-induced apoptosis is associated with PARP leavage, activation of caspase-3, -8, and -9 in a time- and dose-dependent manner. Hence, both caspase-8 and -9-initiated apoptotic signaling pathways contribute to metforrnin-induced apoptosis in pancreatic cell lines.

  6. MG132 Induced Apoptosis Pathway in HL-60 Cells and Impact of Allogeneic Mixed Lymphocyte Reaction

    Institute of Scientific and Technical Information of China (English)

    Yong-ming Zhou; Wei Guo; Hao Zhou; Jin-hua Zhang; Zhi-ping Liu; Mei-xia Yu

    2009-01-01

    Objective: To investigate the proteasome inhibitor MG132-induced apoptosis pathway in HL-60 cells and the role of allogeneic mixed lymphocyte reaction.Methods: Cell apoptosis was analyzed by flow cytometry. The expressions of p21 protein, p27 protein and p53 protein in HL-60 cells treated with MG132 were measured by Western blot. The proliferation of, peripheral blood mononuclear cells (PBMNCs) after treatment with 75 Gy irradiated HL-60 cells treated with MG132 was measured with CCK-8.Results: High-dose MG132 induced apoptosis in HL-60 cells. No significant change was observed in MG132-induced apoptosis after inhibiting caspase-8 and caspase-9 pathway. The expressions of p21 protein and p27 protein increased in MG132-induced apoptosis. HL-60 cells treated with low-dose MG132 improved the proliferation of PBMNCs from healthy volunteers.Conclusion: High-dose MG132 induced apoptosis and directly killed HL-60 cells. MG132 induced apoptosis in a caspase-8- and caspase-9-independent pathway. p21 protein and p27 protein were involved in MG132-induced apoptosis in HL-60 cells. HL-60 cells treated with Low-dose MG132 improved the effect of promoting the proliferation of PBMNCs from healthy volunteers.

  7. Apoptotic pathway induced by noscapine in human myelogenous leukemic cells.

    Science.gov (United States)

    Heidari, Nastaran; Goliaei, Bahram; Moghaddam, Parvaneh Rahimi; Rahbar-Roshandel, Nahid; Mahmoudian, Massoud

    2007-11-01

    It has been shown that noscapine, an opium-derived phthalideisoquinoline alkaloid that is currently being used as an oral antitussive drug, induces apoptosis in myeloid leukemia cells. The molecular mechanism responsible for the anticancer effects of noscapine is poorly understood. In the current study, the apoptotic effects of noscapine on two myeloid cell lines, apoptosis-proficient HL60 cells and apoptosis-resistant K562 cells, were analyzed. An increase in the activity of caspase-2, -3, -6, -8 and -9, poly(ADP ribose) polymerase cleavage, detection of phosphatidylserine on the outer layer of the cell membrane, nucleation of chromatin, and DNA fragmentation suggested the induction of apoptosis. Noscapine increased the Bax/Bcl-2 ratio with a significant decrease of Bcl-2 expression accompanied with Bcl-2 phosphorylation. Using an inhibitory approach, the activation of the caspase cascade involved in the noscapine-induced apoptosis was analyzed. We observed no inhibitory effect of the caspase-8 inhibitor on caspase-9 activity. In view of these results and taking into consideration that K562 cells are Fas-null, we suggested that caspase-8 is activated in a Fas-independent manner downstream of caspase-9. In conclusion, noscapine can induce apoptosis in both apoptosis-proficient and apoptosis-resistant leukemic cells, and it can be a novel candidate in the treatment of hematological malignancies.

  8. Restoration of APC gene function in colorectal cancer cells by aminoglycoside- and macrolide-induced read-through of premature termination codons.

    Science.gov (United States)

    Zilberberg, Alona; Lahav, Lital; Rosin-Arbesfeld, Rina

    2010-04-01

    Adenomatous polyposis coli (APC) is a multifunctional tumour suppressor protein that negatively regulates the Wnt signalling pathway. The APC gene is ubiquitously expressed in tissues and organs, including the large intestine and central nervous system. The majority of patients with sporadic and hereditary colorectal cancer have mutations in the gene encoding APC. Approximately 30% of these mutations are single nucleotide changes that result in premature stop codons (nonsense mutations). A potential therapeutic approach for treatment of this subset of patients is the use of aminoglycosides and macrolides that induce nonsense mutation read-through and restore levels of full-length protein. We have used reporter plasmids and colorectal cancer cell lines to demonstrate that several aminoglycosides and tylosin, a member of the macrolide family, induced read-through of nonsense mutations in the APC gene. In xenograft experiments and in the Apc(Min/+) mouse model, these compounds ameliorated the tumorigenic clinical symptoms caused by nonsense mutations in the APC gene.

  9. In vivo evidence against clomethiazole being neuroprotective against MDMA ('ecstasy')-induced degeneration of rat brain 5-HT nerve terminals by a free radical scavenging mechanism.

    Science.gov (United States)

    Colado, M I; O'Shea, E; Esteban, B; Granados, R; Green, A R

    1999-02-01

    Clomethiazole is an effective neuroprotective agent against the degeneration of 5-HT neurones that follows administration of 3,4-methylenedioxymethamphetamine (MDMA or 'ecstasy'). Since there is good evidence that free radical formation resulting from auto-oxidation of MDMA metabolites is responsible for the degeneration we have examined whether clomethiazole is a free radical scavenger. MDMA (15 mg/kg i.p.) increased the formation of 2,3- and 2,5-dihydroxybenzoic acids (2,3-DHBA and 2,5-DHBA) from salicylic acid perfused through a microdialysis tube implanted in the hippocampus, indicating increased free radical formation. Clomethiazole (50 mg/kg i.p.) administered 5 min prior and 55 min post MDMA prevented both the acute MDMA-induced hyperthermia and the rise in 2,3- and 2,5-DHBA. However, when the temperature of the MDMA + clomethiazole treated rats was kept elevated to that of the MDMA treated rats with a homeothermic blanket there was no inhibition of the MDMA-induced increase in 2,3-DHBA or 2,5-DHBA. These data suggest firstly that free radical formation is inhibited when the acute MDMA-induced hyperthermia is prevented. Secondly the data further indicate that clomethiazole has no free radical scavenging activity since the drug produces substantial neuroprotection when MDMA + clomethiazole treated rats are kept hyperthermic. This conclusion was strengthened by our observation that clomethiazole is a weak inhibitor (IC50 > 1 mM) of lipid peroxidation in synaptosomes when it had been induced by addition of FeCl2 + ascorbic acid.

  10. Hydrogen-Rich Saline Attenuates Lipopolysaccharide-Induced Heart Dysfunction by Restoring Fatty Acid Oxidation in Rats by Mitigating C-Jun N-Terminal Kinase Activation.

    Science.gov (United States)

    Tao, Bingdong; Liu, Lidan; Wang, Ni; Tong, Dongyi; Wang, Wei; Zhang, Jin

    2015-12-01

    Sepsis is common in intensive care units (ICU) and is associated with high mortality. Cardiac dysfunction complicating sepsis is one of the most important causes of this mortality. This dysfunction is due to myocardial inflammation and reduced production of energy by the heart. A number of studies have shown that hydrogen-rich saline (HRS) has a beneficial effect on sepsis. Therefore, we tested whether HRS prevents cardiac dysfunction by increasing cardiac energy. Four groups of rats received intraperitoneal injections of one of the following solutions: normal saline (NS), HRS, lipopolysaccharide (LPS), and LPS plus HRS. Cardiac function was measured by echocardiography 8 h after the injections. Gene and protein expression related to fatty acid oxidation (FAO) were measured by quantitative polymerase chain reaction (PCR) and Western blot analysis. The injection of LPS compromised heart function through decreased fractional shortening (FS) and increased left ventricular diameter (LVD). The addition of HRS increased FS, palmitate triphosphate, and the ratio of phosphocreatinine (PCr) to adenosine triphosphate (ATP) as well as decreasing LVD. The LPS challenge reduced the expression of genes related to FAO, including perioxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), perioxisome proliferator-activated receptor alpha (PPARα), Estrogen-related receptor alpha (ERRα), and their downstream targets, in mRNA and protein level, which were attenuated by HRS. However, HRS had little effect on glucose metabolism. Furthermore, HRS inhibited c-Jun N-terminal kinase (JNK) activation in the rat heart. Inhibition of JNK by HRS showed beneficial effects on LPS-challenged rats, at least in part, by restoring cardiac FAO.

  11. Low Dose Acetaminophen Induces Reversible Mitochondrial Dysfunction Associated with Transient c-Jun N-Terminal Kinase Activation in Mouse Liver.

    Science.gov (United States)

    Hu, Jiangting; Ramshesh, Venkat K; McGill, Mitchell R; Jaeschke, Hartmut; Lemasters, John J

    2016-03-01

    Acetaminophen (APAP) overdose causes hepatotoxicity involving mitochondrial dysfunction and c-jun N-terminal kinase (JNK) activation. Because the safe limit of APAP dosing is controversial, our aim was to evaluate the role of the mitochondrial permeability transition (MPT) and JNK in mitochondrial dysfunction after APAP dosing considered nontoxic by criteria of serum alanine aminotransferase (ALT) release and histological necrosis in vivo. C57BL/6 mice were given APAP with and without the MPT inhibitor, N-methyl-4-isoleucine cyclosporin (NIM811), or the JNK inhibitor, SP600125. Fat droplet formation, cell viability, and mitochondrial function in vivo were monitored by intravital multiphoton microscopy. Serum ALT, liver histology, total JNK, and activated phospho(p)JNK were also assessed. High APAP (300 mg/kg) caused ALT release, necrosis, irreversible mitochondrial dysfunction, and hepatocellular death. By contrast, lower APAP (150 mg/kg) caused reversible mitochondrial dysfunction and fat droplet formation in hepatocytes without ALT release or necrosis. Mitochondrial protein N-acetyl-p-benzoquinone imine adducts correlated with early JNK activation, but irreversible mitochondrial depolarization and necrosis at high dose were associated with sustained JNK activation and translocation to mitochondria. NIM811 prevented cell death and/or mitochondrial depolarization after both high and low dose APAP. After low dose, SP600125 decreased mitochondrial depolarization. In conclusion, low dose APAP produces reversible MPT-dependent mitochondrial dysfunction and steatosis in hepatocytes without causing ALT release or necrosis, whereas high dose leads to irreversible mitochondrial dysfunction and cell death associated with sustained JNK activation. Thus, nontoxic APAP has the potential to cause transient mitochondrial dysfunction that may synergize with other stresses to promote liver damage and steatosis.

  12. Increase of RhoB in {gamma}-radiation-induced apoptosis is regulated by c-Jun N-terminal kinase in Jurkat T cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Chun-Ho [Laboratory of Cytogenetics and Tissue Regeneration, KIRAMS, Seoul 139-706 (Korea, Republic of); Won, Misun; Choi, Chung-Hae; Ahn, Jiwon; Kim, Bo-Kyung [Genome Research Center, KRIBB, Daejeon 305-806 (Korea, Republic of); Song, Kyung-Bin [Department of Food Science and Technology, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Kang, Chang-Mo, E-mail: kangcm@kcch.re.kr [Laboratory of Cytogenetics and Tissue Regeneration, KIRAMS, Seoul 139-706 (Korea, Republic of); Chung, Kyung-Sook, E-mail: kschung@kribb.re.kr [Genome Research Center, KRIBB, Daejeon 305-806 (Korea, Republic of)

    2010-01-08

    The Ras-related small GTP-binding protein RhoB is known to be a pro-apoptotic protein and immediate-early inducible by genotoxic stresses. In addition, JNK activation is known to function in {gamma}-radiation-induced apoptosis. However, it is unclear how JNK activation and {gamma}-radiation-dependent RhoB induction are related. Here we verified the relationship between JNK activation and RhoB induction. RhoB induction by {gamma}-radiation occurred at the transcriptional level and transcriptional activation of RhoB was concomitant with an increase in RhoB protein. {gamma}-Radiation-induced RhoB expression was markedly attenuated by pretreatment with a JNK-specific inhibitor, SP600125, but not by a p38 MAPK inhibitor, SB203580. Inhibition of JNK caused a decrease in early apoptotic cell death that correlated with RhoB expression. However, PI3K inhibition had no significant effects, indicating that the AKT survival pathway was not involved. The siRNA knockdown of JNK resulted in a decrease in RhoB expression and the siRNA knockdown of RhoB restored cell growth even in the {gamma}-irradiated cells. These results suggest that RhoB regulation involves the JNK pathway and contributes to the early apoptotic response of Jurkat T cells to {gamma}-radiation.

  13. Terminated Multifamily Mortgages Database

    Data.gov (United States)

    Department of Housing and Urban Development — This dataset includes all terminated HUD Multifamily mortgages except those from the Hospital Mortgage Insurance Program. It includes the Holder and Servicer at the...

  14. Coal terminal directory

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2008-06-15

    The directory gives a comprehensive listing of the world's coal terminals, in a total of 50 countries including information on throughput, facilities, storage capacity, and vessel size limitation.

  15. Role of α-toxin-induced apoptosis of umbilical vein endothelial cells in vertical infection of Staphylococcus aureus L-form%α-毒素诱导的脐静脉内皮细胞凋亡在金葡菌L型垂直感染中的作用

    Institute of Scientific and Technical Information of China (English)

    管俊昌; 朱翔; 余峰玲; 杨文选; 刘婷婷; 张涛; 林娜; 刘勇; 刘从森

    2013-01-01

    目的 确定α-毒素诱导脐静脉内皮细胞(HUV-EC-C)的凋亡在金葡菌L型垂直感染中的作用.方法 在培养的HUV-EC-C细胞中加入不同浓度(0、10、30、90、270 ng/ml)的金葡菌α-毒素,处理不同时间(0、2、4、6、8h)后经Annexin V-PI染色,流式细胞仪检测HUV-EC-C细胞的凋亡率.通过ELISA检测α肿瘤坏死因子(TNF-α)、caspase-3与caspase-8的表达量,并观察加入TNF-α中口抗体、caspases-3抑制剂zDEVD-FMK、caspase-8抑制剂zIETD-fmk对α-毒素诱导HUV-EC-C细胞凋亡的影响.结果 α-毒素能够诱导HUV-EC-C细胞凋亡并表现为时间、剂量依赖性,明显增加HUV-EC-C细胞中TNF-α、caspase-3与caspase-8的表达;在加入TNF-α中和抗体、zDEVD-FMK、zIETD-fmk后可部分阻断α-毒素能够诱导的HUV-EC-C细胞凋亡.结论 α-毒素通过TNF-α及caspase-8介导的外源性死亡途径诱导HUV-EC-C细胞凋亡,表明α-毒素诱导内皮细胞凋亡是金葡菌L型垂直感染影响胎儿发育的主要机制之一.%Objective To investigate α-toxin-induced apoptosis of umbilical vein endothelial cells and explore its role in vertical infection of Staphylococcus aureus L-form.Methods HUV-EC-C cells exposed to different concentrations (0,10,30,90,and 270 ng/ml) of α-toxin for different time lengths (0,2,4,6,and 8 h) were examined for apoptosis using flow cytometry with Annexin V-PI staining.The levels of tumor necrosis factor-α (TNF-α) and the activities of,caspase-3 and caspase-8 in the cell culture were detected by ELISA and colorimetric method,respectively.α-Toxin-induced cell apoptosis was also analyzed in HUV-EC-C cells treated with a neutralizing antibody of TNF-α or with the inhibitory peptides of caspase-3 (zDEVD-FMK) and caspase-8 (zIETD-fmk).Results α-Toxin induced apoptosis of HUV-EC-C cells in a dose-and time-dependent manner and caused significantly enhanced expression of TNF-α and the activation of both caspase-3 and caspase-8.Inhibition of TNF-α with

  16. Rabbit IgG directed to a synthetic C-terminal peptide of the major grass pollen allergen Lol p I inhibits human basophil histamine release induced by natural Lol p I.

    Science.gov (United States)

    van Ree, R; Aalberse, R C

    1995-03-01

    The potential role of allergen-specific IgG antibodies as 'blocking' antibodies in allergen-induced human basophil histamine release was investigated. This was studied in a model with the major grass pollen allergen Lol p I and polyclonal rabbit antisera directed against this allergen and against a synthetic peptide of its C terminus. When allergen and antibodies were allowed to preincubate, Lol p I induced histamine release was inhibited up to 85% by the antiserum against Lol p I. By omitting preincubation, and thereby more closely mimicking an in vivo situation, up to 55% inhibition was realized. This indicates that allergen-specific IgG can act as 'blocking' antibody without preincubation. Immunization of rabbits with a synthetic C-terminal peptide of Lol p I resulted in antibodies reactive with natural Lol p I. Despite their 100-fold lower avidity for Lol p I (as compared with antinatural Lol p I), these antibodies had the capacity to inhibit Lol p I induced histamine release for > 90% (up to 50% without preincubation). This indicates that it is possible to block histamine release induced by a major allergen with low-avidity IgG antibodies directed against a minor proportion of the allergen (25 amino acids). IgE antibodies from the donors studied were unreactive with this synthetic peptide, indicating that for blocking activity identical epitope specificity of IgE and IgG is not essential. This opens interesting perspectives for application of synthetic peptides in immunotherapy, distinct from their effects on T cell reactivity.

  17. The methylotrophic yeast Hansenula polymorpha contains an inducible import pathway for peroxisomal matrix proteins with an N-terminal targeting signal (PTS2 proteins).

    Science.gov (United States)

    Faber, K N; Haima, P; Gietl, C; Harder, W; Ab, G; Veenhuis, M

    1994-12-20

    Two main types of peroxisomal targeting signals have been identified that reside either at the extreme C terminus (PTS1) or the N terminus (PTS2) of the protein. In the methylotrophic yeast Hansenula polymorpha the majority of peroxisomal matrix proteins are of the PTS1 type. Thus far, for H. polymorpha only amine oxidase (AMO) has been shown to contain a PTS2 type signal. In the present study we expressed H. polymorpha AMO under control of the strong endogenous alcohol oxidase promoter. Partial import of AMO into peroxisomes was observed in cells grown in methanol/(NH4)2SO4-containing medium. However, complete import of AMO occurred if the cells were grown under conditions that induce expression of the endogenous AMO gene. Similar results were obtained when the heterologous PTS2 proteins, glyoxysomal malate dehydrogenase from watermelon and thiolase from Saccharomyces cerevisiae, were synthesized in H. polymorpha. The import of PTS1 proteins, however, was not affected by the growth conditions. These results indicate that the reduced rate of AMO import in (NH4)2SO4-grown cells is not due to competition with PTS1 proteins for the same import pathway. Apparently, AMO is imported via a separate pathway that is induced by amines and functions for PTS2 proteins in general.

  18. Neem oil limonoids induces p53-independent apoptosis and autophagy.

    Science.gov (United States)

    Srivastava, Pragya; Yadav, Neelu; Lella, Ravi; Schneider, Andrea; Jones, Anthony; Marlowe, Timothy; Lovett, Gabrielle; O'Loughlin, Kieran; Minderman, Hans; Gogada, Raghu; Chandra, Dhyan

    2012-11-01

    Azadirachta indica, commonly known as neem, has a wide range of medicinal properties. Neem extracts and its purified products have been examined for induction of apoptosis in multiple cancer cell types; however, its underlying mechanisms remain undefined. We show that neem oil (i.e., neem), which contains majority of neem limonoids including azadirachtin, induced apoptotic and autophagic cell death. Gene silencing demonstrated that caspase cascade was initiated by the activation of caspase-9, whereas caspase-8 was also activated late during neem-induced apoptosis. Pretreatment of cancer cells with pan caspase inhibitor, z-VAD inhibited activities of both initiator caspases (e.g., caspase-8 and -9) and executioner caspase-3. Neem induced the release of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria, suggesting the involvement of both caspase-dependent and AIF-mediated apoptosis. p21 deficiency caused an increase in caspase activities at lower doses of neem, whereas p53 deficiency did not modulate neem-induced caspase activation. Additionally, neem treatment resulted in the accumulation of LC3-II in cancer cells, suggesting the involvement of autophagy in neem-induced cancer cell death. Low doses of autophagy inhibitors (i.e., 3-methyladenine and LY294002) did not prevent accumulation of neem-induced LC3-II in cancer cells. Silencing of ATG5 or Beclin-1 further enhanced neem-induced cell death. Phosphoinositide 3-kinase (PI3K) or autophagy inhibitors increased neem-induced caspase-3 activation and inhibition of caspases enhanced neem-induced autophagy. Together, for the first time, we demonstrate that neem induces caspase-dependent and AIF-mediated apoptosis, and autophagy in cancer cells.

  19. Documents from malicious terminals

    Science.gov (United States)

    Berta, Istvan Z.; Vajda, Istvan

    2003-04-01

    The user wishes to communicate with a remote partner over an insecure network. Since the user is a human being, a terminal is needed for communication. Cryptographic algorithms running on the terminal may provide authenticity for the user's messages. In this paper the problem of sending authentic messages from insecure or untrusted terminals is analyzed. In this case attackers are able to gain total control over the terminal, so the user must consider the terminal a potential attacker. Smart cards are often considered the ultimate tool for secure messaging from untrusted terminals. However, their lack of user interface enables man-in-the middle attack from the terminal. The authors assume, that user is a human being with limited memory and computational power, and also makes mistakes in his calculations. They demnostrate, that only exceptional useres are able to authenticate messages without a trusted device. Several biometric media encapsulate the content of the message and the identity of the sender, such as speech, video and handwriting. The authors suggest, that such media is far more difficult to counterfeit than plaintext. Thus, the user must rely on his other resources, like biometric ones. In the protocol proposed by the authors, the user sends messages in a biometric format, strengthened by simple algorithmic authenticators. The smart card functions as a secure time gate ensuring, that the attacker has extremely little time to counterfeit both the biometric and the algorithmic protection on the message. The authors claim, that with the proper calibration of the biometric method and the time gate of the smart card, their protocol is strong enough for practical use.

  20. Local Termination: theory and practice

    CERN Document Server

    Endrullis, Joerg; Waldmann, Johannes

    2010-01-01

    The characterisation of termination using well-founded monotone algebras has been a milestone on the way to automated termination techniques, of which we have seen an extensive development over the past years. Both the semantic characterisation and most known termination methods are concerned with global termination, uniformly of all the terms of a term rewriting system (TRS). In this paper we consider local termination, of specific sets of terms within a given TRS. The principal goal of this paper is generalising the semantic characterisation of global termination to local termination. This is made possible by admitting the well-founded monotone algebras to be partial. We also extend our approach to local relative termination. The interest in local termination naturally arises in program verification, where one is probably interested only in sensible inputs, or just wants to characterise the set of inputs for which a program terminates. Local termination will be also be of interest when dealing with a specif...

  1. 药物与人工流产终止早期妊娠的临床比较%Clinical comparison on termination of early pregnancy with medical abortion and induced abortion

    Institute of Scientific and Technical Information of China (English)

    柯梅英; 岑秋妹; 冯亦芳

    2014-01-01

    目的:比较药物流产和人工流产两种不同方式在终止早期妊娠上的被接受程度以及临床效果。方法选取2008~2012年在茂名市人口和计划生育综合服务中心终止早期妊娠的286例受术者,根据终止妊娠方式的不同,分为药物流产组(128例)和人工流产组(158例),比较两组受术者终止妊娠的临床效果以及再次意外妊娠后会选择的终止妊娠方式。结果两组在完全流产率、不完全流产率、人工流产综合征发生率、流产后阴道出血时间上比较差异无统计学意义(P >0.05)。两组再次意外妊娠后会选择的终止妊娠方式比较,差异有统计学意义(P 0. 05). Significant difference were fOund between twO grOuPs in chOOsing the terminatiOn methOd Of Pregnancy with anOther unexPected Pregnancy(P < 0. 05). Conclusion Drug abOrtiOn and induced abOrtiOn in the terminatiOn Of early Pregnancy have different advantages and disadvantages,aPPrOPriate terminatiOn Of early Pregnancy shOuld be chOsen accOrding tO the Patientˊs sPecific situatiOn and PreviOus abOrtiOn histOry tO imPrOve the clinical effect.

  2. c-Jun N-terminal kinase inhibitor favors transforming growth factor-β to antagonize hepatitis B virus X protein-induced cell growth promotion in hepatocellular carcinoma.

    Science.gov (United States)

    Wu, Yan-Hui; Ai, Xi; Liu, Fu-Yao; Liang, Hui-Fang; Zhang, Bi-Xiang; Chen, Xiao-Ping

    2016-02-01

    Transforming growth factor (TGF)-β induces cell growth arrest in well-differentiated hepatocellular carcinoma (HCC) while hepatitis B virus X protein (HBx) minimizes the tumor suppression of TGF-β signaling in early chronic hepatitis B. However, how to reverse the oncogenic effect of HBx and sustain the tumor-suppressive action of TGF-β has yet to be investigated. The present study examined the effect of TGF-β and a c-Jun N-terminal kinase (JNK) inhibitor on cell growth in HCC cells with forced expression of HBx. It was found that HBx promoted cell growth via activation of the JNK/pSMAD3L pathway and inhibition of the transforming growth factor-beta type I receptor (TβRI)/pSMAD3C pathway. pSMAD3L/SMAD4 and pSMAD3C/SMAD4 complexes antagonized each other to regulate c-Myc expression. In the absence of HBx, TGF-β induced cell growth arrest through activation of the TβRI/pSMAD3C pathway in well-differentiated HCC cells. In the presence of HBx, TGF-β had no effect on cell growth. JNK inhibitor SP600125 significantly reversed the oncogenic action of HBx and favored TGF-β to regain the ability to inhibit the cell growth in HBx-expressing well-differentiated HCC cells. In conclusion, targeting JNK signaling favors TGF-β to block HBx-induced cell growth promotion in well-differentiated HCC cells. As an adjunct to anti-viral therapy, the combination of TGF-β and inhibition of JNK signaling is a potential therapy for HBV-infected HCC.

  3. Prenatal alcohol exposure inducing the apoptosis of mossy cells in hippocampus of SMS2-/- mice.

    Science.gov (United States)

    Wang, Lai; Wu, Lin; Wang, Xiaoqing; Deng, Jiexin; Ma, Zhanyou; Fan, Wenjuan; He, Weiya; Deng, Jinbo

    2015-11-01

    In order to understand the mechanisms of alcohol-induced neuroapoptosis through the ceramide pathway, sphingomyelin synthase 2 knockout (SMS2-/-) mice were used to make the prenatal alcohol exposure model, and the role of ceramide regulation on alcohol-induced neuroapoptosis was studied in the offspring. Initially the levels of serum sphingomyelin (SM) were detected with enzymatic method in P0 pups after alcohol exposure in parents. Then the apoptosis of mossy cells in the offspring hippocampus was investigated after prenatal alcohol exposure with immunohistochemistry and TUNEL assay. Finally the expression of activated Caspase 8 and activated Caspase 3 in the offspring hippocampus was detected with Western blot analysis. Our results showed that SM levels were down-regulated in a dose-dependent manner (palcohol exposure in wild-type (WT) and SMS2-/- pups. However, SM levels of serum in SMS2-/- pups were significantly lower than that in WT pups (palcohol-induced neuroapoptosis. In both WT pups and SMS2-/- pups, the number of apoptotic mossy cells in the hippocampus increased after prenatal alcohol exposure in a dose dependent manner (palcohol exposure, consistent with results from TUNEL assay and immunocytochemistry. Our study suggests that mossy cells may be the easily attacked cells for fetal alcohol spectrum disorder (FASD), and ceramide is involved in the alcohol-induced neural apoptosis. The mechanism probably lies in the accumulated ceramide in SMS2 mice, and the increase of activated Caspase 8 and Caspase 3 promotes alcohol-induced neuroapoptosis.

  4. XIAP acts as a switch between type I and type II FAS-induced apoptosis signalling

    OpenAIRE

    Jost, Philipp J; Grabow, Stephanie; Gray, Daniel; McKenzie, Mark D.; Nachbur, Ueli; Huang, David C. S.; Bouillet, Philippe; Thomas, Helen E.; Borner, Christoph; Silke, John; Strasser, Andreas; Kaufmann, Thomas

    2009-01-01

    FAS (APO-1/CD95) and its physiological ligand, FASL, regulate apoptotic death of unwanted or dangerous cells in many tissues, functioning as a guardian against autoimmunity and cancer development1-4. Distinct cell types differ in the mechanisms by which the ‘death receptor’ FAS triggers their apoptosis1-4. In type I cells, such as lymphocytes, activation of ‘effector caspases’ by FAS-induced activation of caspase-8 suffices for cell killing whereas in type II cells, including hepatocytes and ...

  5. Downregulation of miR-181b in mouse brain following ischemic stroke induces neuroprotection against ischemic injury through targeting heat shock protein A5 and ubiquitin carboxyl-terminal hydrolase isozyme L1.

    Science.gov (United States)

    Peng, Zhifeng; Li, Jiefei; Li, Yun; Yang, Xuan; Feng, Sujuan; Han, Song; Li, Junfa

    2013-10-01

    Understanding the molecular mechanism of cerebral hypoxic preconditioning (HPC)-induced endogenous neuroprotection may provide potential therapeutic targets for ischemic stroke. By using bioinformatics analysis, we found that miR-181b, one of 19 differentially expressed miRNAs, may target aconitate hydratase (ACO2), heat shock protein A5 (HSPA5), and ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCHL1) among 26 changed protein kinase C isoform-specific interacting proteins in HPC mouse brain. In this study, the role of miR-181b in oxygen-glucose deprivation (OGD)-induced N2A cell ischemic injury in vitro and mouse middle cerebral artery occlusion (MCAO)-induced cerebral ischemic injury in vivo, and its regulation of ACO2, HSPA5, and UCHL1 were further determined. We found that miR-181b expression levels significantly decreased in mouse brain following MCAO and in OGD-treated N2A cells. Up- and downregulation of miR-181b by transfection of pre- or anti-miR-181b could negatively regulate HSPA5 and UCHL1 (but not ACO2) protein levels as well as N2A cell death and programmed cell death in OGD-treated N2A cells. By using a T7 promoter-driven control dual luciferase assay, we confirmed that miR-181b could bind to the 3'-untranslated rergions of HSPA5 and UCHL1 mRNAs and repress their translations. miR-181b antagomir reduced caspase-3 cleavage and neural cell loss in cerebral ischemic cortex and improved neurological deficit of mice after MCAO. In addition, HSPA5 and UCHL1 short interfering RNAs (siRNAs) blocked anti-miR-181b-mediated neuroprotection against OGD-induced N2A cell injury in vitro. These results suggest that the downregulated miR-181b induces neuroprotection against ischemic injury through negatively regulating HSPA5 and UCHL1 protein levels, providing a potential therapeutic target for ischemic stroke.

  6. Aspartame-induced apoptosis in PC12 cells.

    Science.gov (United States)

    Horio, Yukari; Sun, Yongkun; Liu, Chuang; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    Aspartame is an artificial sweetner added to many low-calorie foods. The safety of aspartame remains controversial even though there are many studies on its risks. In this study, to understand the physiological effects of trace amounts of artificial sweetners on cells, the effects of aspartame on apoptosis were investigated using a PC12 cell system. In addition, the mechanism of apoptosis induced by aspartame in PC12 cells and effects on apoptotic factors such as cytochrome c, apoptosis-inducing factor, and caspase family proteins were studied by Western blotting and RT-PCR. Aspartame-induced apoptosis in PC12 cells in a dose-dependent manner. In addition, aspartame exposure increased the expressions of caspases 8 and 9, and cytochrome c. These results indicate that aspartame induces apoptosis mainly via mitochondrial pathway involved in apoptosis due to oxigen toxicity.

  7. TNF Accelerates Death of Mandibular Condyle Chondrocytes in Rats with Biomechanical Stimulation-Induced Temporomandibular Joint Disease

    Science.gov (United States)

    Zhang, Hongyun; Zhang, Jing; Jing, Lei; Liao, Lifan; Wang, Meiqing

    2015-01-01

    Objective To determine if temporomandibular joint chondrocyte apoptosis is induced in rats with dental biomechanical stimulation and what a role TNF takes. Methods Thirty-two rats were divided into 4 groups (n = 8/group) and exposed to incisor mal-occlusion induced by unilateral anterior crossbite biomechanical stimulation. Two groups were sampled at 2 or 4 weeks. The other two groups were treated with local injections of a TNF inhibitor or PBS into the temporomandibular joints area at 2 weeks and then sampled at 4 weeks. Twenty-four rats either served as unilateral anterior crossbite mock operation controls (n = 8/group) with sampling at 2 or 4 weeks or received a local injection of the TNF inhibitor at 2 weeks with sampling at 4 weeks. Chondrocytes were isolated from the temporomandibular joints of 6 additional rats and treated with TNF in vitro. Joint samples were assessed using Hematoxylin&eosin, Safranin O, TUNEL and immunohistochemistry staining, real-time PCR, fluorogenic activity assays and Western blot analyses. The isolated chondrocytes were also analyzed by flow cytometry. Results Unilateral anterior crossbite stimulation led to temporomandibular joint cartilage degradation, associated with an increase in TUNEL-positive chondrocytes number, caspase-9 expression levels, and the release of cytochrome c from mitochondria at 2 weeks without changes in TNF and caspase-8 levels until after 4 weeks. TNF stimulated apoptosis of the isolated chondrocytes and up-regulated caspase-8 expression, but did not change caspase-9 expression levels. Local injection of TNF inhibitor down-regulated caspase-8 expression and reduced TUNEL-positive cell number, but did not reverse cartilage thickness reduction, caspase-9 up-regulation or cytochrome c release. Conclusions Unilateral anterior crossbite stimulation induces mitochondrion-mediated apoptosis of articular chondrocytes. TNF accelerated the unilateral anterior crossbite induced chondrocytes apoptosis via death

  8. TNF Accelerates Death of Mandibular Condyle Chondrocytes in Rats with Biomechanical Stimulation-Induced Temporomandibular Joint Disease.

    Directory of Open Access Journals (Sweden)

    Hongxu Yang

    Full Text Available To determine if temporomandibular joint chondrocyte apoptosis is induced in rats with dental biomechanical stimulation and what a role TNF takes.Thirty-two rats were divided into 4 groups (n = 8/group and exposed to incisor mal-occlusion induced by unilateral anterior crossbite biomechanical stimulation. Two groups were sampled at 2 or 4 weeks. The other two groups were treated with local injections of a TNF inhibitor or PBS into the temporomandibular joints area at 2 weeks and then sampled at 4 weeks. Twenty-four rats either served as unilateral anterior crossbite mock operation controls (n = 8/group with sampling at 2 or 4 weeks or received a local injection of the TNF inhibitor at 2 weeks with sampling at 4 weeks. Chondrocytes were isolated from the temporomandibular joints of 6 additional rats and treated with TNF in vitro. Joint samples were assessed using Hematoxylin&eosin, Safranin O, TUNEL and immunohistochemistry staining, real-time PCR, fluorogenic activity assays and Western blot analyses. The isolated chondrocytes were also analyzed by flow cytometry.Unilateral anterior crossbite stimulation led to temporomandibular joint cartilage degradation, associated with an increase in TUNEL-positive chondrocytes number, caspase-9 expression levels, and the release of cytochrome c from mitochondria at 2 weeks without changes in TNF and caspase-8 levels until after 4 weeks. TNF stimulated apoptosis of the isolated chondrocytes and up-regulated caspase-8 expression, but did not change caspase-9 expression levels. Local injection of TNF inhibitor down-regulated caspase-8 expression and reduced TUNEL-positive cell number, but did not reverse cartilage thickness reduction, caspase-9 up-regulation or cytochrome c release.Unilateral anterior crossbite stimulation induces mitochondrion-mediated apoptosis of articular chondrocytes. TNF accelerated the unilateral anterior crossbite induced chondrocytes apoptosis via death-receptor pathway. However, anti

  9. Hepatitis B Virus X Protein Sensitizes TRAIL-Induced Hepatocyte Apoptosis by Inhibiting the E3 Ubiquitin Ligase A20.

    Directory of Open Access Journals (Sweden)

    Hang Zhang

    Full Text Available Hepatitis B virus (HBV infection causes hepatocyte death and liver damage, which may eventually lead to cirrhosis and liver cancer. Hepatitis B virus X protein (HBx is a key antigen that is critically involved in HBV-associated liver diseases. However, the molecular basis for its pathogenesis, particularly in liver damage, has not been well defined. Herein, we report that HBx was able to enhance the susceptibility of hepatocytes to TNF-related apoptosis-inducing ligand (TRAIL-induced apoptosis. Increased sensitivity to TRAIL was associated with HBx-induced upregulation of miR-125a, which, in turn, suppressed the expression of its putative target gene, A20 E3 ligase. Importantly, we demonstrate that the defective expression of A20 impaired the K63-linked polyubiquitination of caspase-8, which reciprocally enhanced the activation of caspase-8, the recruitment of Fas-associated death domain (FADD, and the formation of death-inducing signaling complex (DISC, thereby promoting HBx-mediated apoptotic signaling. Accordingly, antagonizing miR-125a or ectopically expressing A20 in hepatocytes abolished the pro-apoptotic effect of HBx. Conversely, the overexpression of miR-125a or knockdown of A20 mimicked HBx to enhance TRAIL susceptibility in hepatocytes. Thus, we establish, for the first time, a miR-125a/A20-initiated and caspase-8-targeted mechanism by which HBx modulates apoptotic signaling and increases hepatic susceptibility to the damaging agent, which might provide novel insight into HBV-related liver pathology.

  10. Effects of Fas, NF-κB and caspases on rat microvascular endothelial cell apoptosis induced by TNFα

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    AIM: To study the apoptotic effect of TNFα on rat pulmonary microvascular endothelial cells (PMVEC) and the influences of Fas, NF-κB in its mechanism. METHODS: Apoptosis of PMVEC was analyzed and quantitated with TUNEL, flow cytometer. The distribution of NF-κB was detected via histoimmunochemical staining in TNF-treated cells and the control. Northern blot was applied to assess the influence of TNF on PMVEC Fas expression. Fas antibody was used to investigate the apoptotic effect of Fas on PMVEC. Activation of caspase-8 was detected with Western blot. Expression of caspase-3 was analyzed with histoimmunochemical staining. RESULTS: After treatment with 5×108 U/L TNF for 24 hours, viable PMVEC significantly diminished. Apoptosis rate was 14.0%±3.1% detected with TUNEL, and 13.1% with flow cytometer. Histoimmunochemical staining showed that NF-κB relocated from cytoplasm to the nuclear. When the cells were co-cultured with TNF and APDC, an NF-κB inhibitor, less cells were viable and more cells were positively stained with TUNEL. Fas expression in PMVEC was elevated treated with TNF. Apoptosis in PMVEC was found aggravated, when the cells were co-cultured with TNF and anti-Fas antibody. The positive rate was 24.1%±1.5% with TUNEL. Increase of caspase-8 activation was manifested by Western blot following TNF stimulation. Caspase-3 expression was found elevated using histoimmunochemical staining. Cell permeable caspase-3 inhibitor significantly ameliorated PMVEC apoptosis induced by TNF. CONCLUSION: 1. Large dose of TNF(5×108 U/L) can induce apoptosis in rat PMVEC. 2. NF-κB has a protective effect on PMVEC apoptosis. 3. TNF up-regulates Fas expression in PMVEC. And the latter takes a part in apoptosis. 4. TNF induced caspase-8 activation in PMVEC, and more caspase-3 was expressed. These may be involved in PMVEC apoptosis induced by TNF.

  11. Modeling Terminal Velocity

    Science.gov (United States)

    Brand, Neal; Quintanilla, John A.

    2013-01-01

    Using a simultaneously falling softball as a stopwatch, the terminal velocity of a whiffle ball can be obtained to surprisingly high accuracy with only common household equipment. This classroom activity engages students in an apparently daunting task that nevertheless is tractable, using a simple model and mathematical techniques at their…

  12. Navy Multiband Terminal (NMT)

    Science.gov (United States)

    2015-12-01

    military satellite communications terminal. The NMT Program is the required Navy component to the Advanced Extremely High Frequency (AEHF) Program... Advanced Extremely High Frequency ATO - Approval to Operate DAA - Designated Approval Authority deg - degree DISR - DoD Information Standards...Intermediate, and Depot. The Intermediate maintenance will be performed by the Regional Maintenance Centers and further supported by the In Service

  13. A B-Cell Superantigen Induces the Apoptosis of Murine and Human Malignant B Cells

    Science.gov (United States)

    Lorenzo, Daniela; Duarte, Alejandra; Mundiñano, Juliana; Berguer, Paula; Nepomnaschy, Irene; Piazzon, Isabel

    2016-01-01

    B-cell superantigens (Sags) bind to conserved sites of the VH or VL regions of immunoglobulin molecules outside their complementarity-determining regions causing the apoptosis of normal cognate B cells. No attempts to investigate whether B-cell Sags are able to induce the apoptosis of cognate malignant B cells were reported. In the present study we show that protein L (PpL), secreted by Finegoldia magna, a B-cell Sag which interacts with κ+ bearing cells, induces the apoptosis of murine and human κ+ lymphoma B cells both in vitro and in vivo. Apoptosis was not altered by caspase-8 inhibitor. No alterations in the levels of Bid, Fas and Fas-L were found suggesting that PpL does not activate the extrinsic pathway of apoptosis. The involvement of the intrinsic pathway was clearly indicated by: i) alterations in mitochondrial membrane potential (ΔΨm) both in murine and human lymphoma cells exposed to PpL; ii) decreased levels of apoptosis in the presence of caspase-9 inhibitor; iii) significant increases of Bim and Bax protein levels and downregulation of Bcl-2; iv) the translocation from the cytoplasm to the mitochondria of Bax and Bim pro-apoptotic proteins and its inhibition by caspase-9 inhibitor but not by caspase-8 inhibitor and v) the translocation of Bcl-2 protein from the mitochondria to the cytosol and its inhibition by caspase-9 inhibitor but not by caspase-8 inhibitor. The possibility of a therapeutic use of Sags in lymphoma/leukemia B cell malignancies is discussed. PMID:27603942

  14. BNip3 is a mediator of TNF-induced necrotic cell death.

    Science.gov (United States)

    Kim, Jee-Youn; Kim, Yong-Jun; Lee, Sun; Park, Jae-Hoon

    2011-02-01

    Tumor necrosis factor (TNF) is a pleiotropic cytokine involved in immune modulation, inflammatory reactions, and target cell death in many pathologic conditions. The cell death pathways triggered by TNF include the caspase-8/Bid-dependent apoptotic pathway and the caspase-independent necrosis pathway (necroptosis). While the signaling pathways activated after binding of TNF to the TNF receptor (TNFR) and subsequent insertion of Bid/Bax/Bik into the outer mitochondrial membrane are relatively well known, other cell death pathways and the participating signaling molecules remain to be clarified. BNip3 is a pro-death protein and a member of the BH3-only Bcl-2 family. When ectopically overexpressed or induced by hypoxia, BNip3 induces various types of cell death via mitochondrial or non-mitochondrial death cascades. In this study using A549 alveolar epithelial cells of the lung, we show that BNip3 is transcriptionally and translationally upregulated by TNF, and its expression level determines the sensitivity to necroptosis induced by TNF. However, BNip3 does not appear to be involved in caspase-8/Bid-dependent apoptotic cell death in these alveolar lung cells. Finally, we show that the generation of reactive oxygen species (ROS) is essential for mitochondrial insertion of BNip3, which is an important step in BNip3-induced mitochondrial catastrophe. Our results indicate that BNip3 is a candidate therapeutic target in pathologic conditions in which TNF causes tissue damage.

  15. Pregnane X Receptor Activation Attenuates Inflammation-Associated Intestinal Epithelial Barrier Dysfunction by Inhibiting Cytokine-Induced Myosin Light-Chain Kinase Expression and c-Jun N-Terminal Kinase 1/2 Activation.

    Science.gov (United States)

    Garg, Aditya; Zhao, Angela; Erickson, Sarah L; Mukherjee, Subhajit; Lau, Aik Jiang; Alston, Laurie; Chang, Thomas K H; Mani, Sridhar; Hirota, Simon A

    2016-10-01

    The inflammatory bowel diseases (IBDs) are chronic inflammatory disorders with a complex etiology. IBD is thought to arise in genetically susceptible individuals in the context of aberrant interactions with the intestinal microbiota and other environmental risk factors. Recently, the pregnane X receptor (PXR) was identified as a sensor for microbial metabolites, whose activation can regulate the intestinal epithelial barrier. Mutations in NR1I2, the gene that encodes the PXR, have been linked to IBD, and in animal models, PXR deletion leads to barrier dysfunction. In the current study, we sought to assess the mechanism(s) through which the PXR regulates barrier function during inflammation. In Caco-2 intestinal epithelial cell monolayers, tumor necrosis factor-α/interferon-γ exposure disrupted the barrier and triggered zonula occludens-1 relocalization, increased expression of myosin light-chain kinase (MLCK), and activation of c-Jun N-terminal kinase 1/2 (JNK1/2). Activation of the PXR [rifaximin and [[3,5-Bis(1,1-dimethylethyl)-4-hydroxyphenyl]ethenylidene]bis-phosphonic acid tetraethyl ester (SR12813); 10 μM] protected the barrier, an effect that was associated with attenuated MLCK expression and JNK1/2 activation. In vivo, activation of the PXR [pregnenolone 16α-carbonitrile (PCN)] attenuated barrier disruption induced by toll-like receptor 4 activation in wild-type, but not Pxr-/-, mice. Furthermore, PCN treatment protected the barrier in the dextran-sulfate sodium model of experimental colitis, an effect that was associated with reduced expression of mucosal MLCK and phosphorylated JNK1/2. Together, our data suggest that the PXR regulates the intestinal epithelial barrier during inflammation by modulating cytokine-induced MLCK expression and JNK1/2 activation. Thus, targeting the PXR may prove beneficial for the treatment of inflammation-associated barrier disruption in the context of IBD.

  16. c-Jun N-terminal kinases 3 (JNK3) from orange-spotted grouper, Epinephelus coioides, inhibiting the replication of Singapore grouper iridovirus (SGIV) and SGIV-induced apoptosis.

    Science.gov (United States)

    Guo, Minglan; Wei, Jingguang; Zhou, Yongcan; Qin, Qiwei

    2016-12-01

    C-Jun N-terminal kinases (JNKs), a subgroup of serine-threonine protein kinases that activated by phosphorylation, are involve in physiological and pathophysiological processes. JNK3 is one of JNK proteins involved in JNK3 signaling transduction. In the present study, two JNK3 isoforms, Ec-JNK3 X1 and Ec-JNK3 X2, were cloned from orange-spotted grouper, Epinephelus coioides. Both Ec-JNK3 X1 and Ec-JNK3 X2 were mainly expressed in liver, gill, skin, brain and muscle of juvenile grouper. The relative expression of Ec-JNK3 X2 mRNA was much higher in muscle and gill than that of Ec-JNK3 X1. Isoform-specific immune response to challenges was revealed by the expression profiles in vivo. Immunofluorescence staining indicated that JNK3 was localized in the cytoplasm of grouper spleen (GS) cells and shown immune response to SGIV infection in vitro. Over-expressing Ec-JNK3 X1 and/or Ec-JNK3 X2 inhibited the SGIV infection and replication and the SGIV-induced apoptosis. To achieve the antiviral and anti-apoptosis activities, JNK3 promoted the activation of genes ISRE and type I IFN in the antiviral IFN signaling pathway, and inhibited the activation of transcription factors NF-κB and p53 relating to apoptosis, respectively. Ec-JNK3 X2 showed stronger activities in antivirus and anti-apoptosis than that of Ec-JNK3 X1. Our results not only define the characterization of JNK3 but also reveal new immune functions and the molecular mechanisms of JNK3 on iridoviruses infection and the virus-induced apoptosis.

  17. Artemisinin induces A549 cell apoptosis dominantly via a reactive oxygen species-mediated amplification activation loop among caspase-9, -8 and -3.

    Science.gov (United States)

    Gao, Weijie; Xiao, Fenglian; Wang, Xiaoping; Chen, Tongsheng

    2013-10-01

    This report is designed to explore the roles of caspase-8, -9 and -3 in artemisinin (ARTE)-induced apoptosis in non-small cell lung cancer cells (A549 cells). ARTE induced reactive oxygen species (ROS)-mediated apoptosis in dose- and time-dependent fashion. Although ARTE treatment did not induce Bid cleavage and significant loss of mitochondrial membrane potential, it induced release of Smac and AIF but not cytochrome c from mitochondria, and silencing of Bak but not Bax significantly prevented ARTE-induced cytotoxicity. Moreover, ARTE treatment induced ROS-dependent activation of caspase-9, -8 and -3. Of the utmost importance, silencing or inhibiting any one of caspase-8, -9 and -3 almost completely prevented ARTE-induced activation of all the three caspases and remarkably abrogated the cytotoxicity of ARTE, suggesting that ARTE triggered an amplification activation loop among caspase-9, -8 and -3. Collectively, our data demonstrate that ARTE induces a ROS-mediated amplification activation loop among caspase-9, -8 and -3 to dominantly mediate the apoptosis of A549 cells.

  18. Cirrus Crystal Terminal Velocities.

    Science.gov (United States)

    Heymsfield, Andrew J.; Iaquinta, Jean

    2000-04-01

    Cirrus crystal terminal velocities are of primary importance in determining the rate of transport of condensate from upper- to middle-tropospheric levels and profoundly influence the earth's radiation balance through their effect on the rate of buildup or decay of cirrus clouds. In this study, laboratory and field-based cirrus crystal drag coefficient data, as well as analytical descriptions of cirrus crystal shapes, are used to derive more physically based expressions for the velocities of cirrus crystals than have been available in the past.Polycrystals-often bullet rosettes-are shown to be the dominant crystal types in synoptically generated cirrus, with columns present in varying but relatively large percentages, depending on the cloud. The two critical parameters needed to calculate terminal velocity are the drag coefficient and the ratio of mass to cross-sectional area normal to their fall direction. Using measurements and calculations, it is shown that drag coefficients from theory and laboratory studies are applicable to crystals of the types found in cirrus. The ratio of the mass to area, which is shown to be relatively independent of the number of bullets in the rosette, is derived from an analytic model that represents bullet rosettes containing one to eight bullets in 19 primary geometric configurations. The ratio is also derived for columns. Using this information, a general set of equations is developed to calculate the terminal velocities and masses in terms of the aspect ratio (width divided by length), ice density, and rosette maximum dimension. Simple expressions for terminal velocity and mass as a function of bullet rosette maximum dimension are developed by incorporating new information on bullet aspect ratios.The general terminal velocity and mass relations are then applied to a case from the First International Satellite Cloud Climatology Project (ISCCP) Research Experiment (FIRE) 2, when size spectra from a balloon-borne ice crystal

  19. Antecedents of Customer Relationship Termination

    DEFF Research Database (Denmark)

    Geersbro, Jens; Ritter, Thomas

    To end business relationships, or to more actively terminate relationships, has long been acknowledged as part of customer relationship management. However, compared to other elements such as initiation and maintenance of relationships, little is known about the termination of business...... relationships as a managerial task. This paper contributes by (1) developing a conceptualization of relationship termination competence and (2) analyzing its antecedents. The empirical results identify termination acceptance, definition non-customers, organizational relationship termination routines......, and motivation as significant antecedents. Because of this, managers need to develop their organizations in order to use relationship termination as a vital strategy....

  20. Termination Casts: A Flexible Approach to Termination with General Recursion

    Directory of Open Access Journals (Sweden)

    Aaron Stump

    2010-12-01

    Full Text Available This paper proposes a type-and-effect system called Teqt, which distinguishes terminating terms and total functions from possibly diverging terms and partial functions, for a lambda calculus with general recursion and equality types. The central idea is to include a primitive type-form "Terminates t", expressing that term t is terminating; and then allow terms t to be coerced from possibly diverging to total, using a proof of Terminates t. We call such coercions termination casts, and show how to implement terminating recursion using them. For the meta-theory of the system, we describe a translation from Teqt to a logical theory of termination for general recursive, simply typed functions. Every typing judgment of Teqt is translated to a theorem expressing the appropriate termination property of the computational part of the Teqt term.

  1. Suramin induces and enhances apoptosis in a model of hyperoxia-induced oligodendrocyte injury.

    Science.gov (United States)

    Stark, Simone; Schuller, Alexandra; Sifringer, Marco; Gerstner, Bettina; Brehmer, Felix; Weber, Sven; Altmann, Rodica; Obladen, Michael; Buhrer, Christoph; Felderhoff-Mueser, Ursula

    2008-01-01

    Recent evidence suggests oxygen as a powerful trigger for cell death in the immature white matter, leading to periventricular leukomalacia (PVL) as a cause of adverse neurological outcome in survivors of preterm birth. This oligodendrocyte (OL) death is associated with oxidative stress, upregulation of apoptotic signaling factors (i.e., Fas, caspase-3) and decreased amounts of neurotrophins. In search of neuroprotective strategies we investigated whether the polysulfonated urea derivative suramin, recently identified as a potent inhibitor of Fas signaling, affords neuroprotection in an in vitro model of hyperoxia-induced injury to immature oligodendrocytes. Immature OLs (OLN-93) were subjected to 80% hyperoxia (48 h) in the presence or absence of suramin (0, 30, 60, 120 microM). Cell death was assessed by flow cytometry (Annexin V, caspase-3 activity assay) and immunohistochemistry for activated caspase-3. Immunoblotting for the death receptor Fas, cleaved caspase-8 and the phosphorylated isoform of the serine-threonin kinase Akt (pAkt) was performed. Suramin lead to OL apoptosis and potentiated hyperoxia-induced injury in a dose-dependent manner. Immunoblotting revealed increased Fas and caspase-8 expression by suramin treatment. This effect was significantly enhanced when suramin was combined with hyperoxia. Furthermore, pAkt levels decreased following suramin exposure, indicating interference with neurotrophin-dependent growth factor signaling. These data indicate that suramin causes apoptotic cell death and aggravates hyperoxia-induced cell death in immature OLs. Its mechanism of action includes an increase of previously described hyperoxia-induced expression of pro-apoptotic factors and deprivation of growth factor dependent signaling components.

  2. Mobile Phone Terminal

    Science.gov (United States)

    1978-01-01

    In the photo, an employee of a real estate firm is contacting his office by means of HICOM, an advanced central terminal for mobile telephones. Developed by the Orlando Division of Martin Marietta Aerospace, Orlando, Florida, and manufactured by Harris Corporation's RF Division, Rochester, N.Y., HICOM upgrades service to users, provides better system management to telephone companies, and makes more efficient use of available mobile telephone channels through a computerized central control terminal. The real estate man, for example, was able to dial his office and he could also have direct-dialed a long distance number. Mobile phones in most areas not yet served by HICOM require an operator's assistance for both local and long distance calls. HICOM improves system management by automatically recording information on all calls for accurate billing, running continual performance checks on its own operation, and reporting any malfunctions to a central office.

  3. Navy Multiband Terminal (NMT)

    Science.gov (United States)

    2013-12-01

    Acquisition Management Information Retrieval Dev Est - Development Estimate DoD - Department of Defense DSN - Defense Switched Network Econ - Economic Eng...Memo Note for Shore (for MTBF and MTBCF): Represents IOT &E and Verification of Correction of Deficiencies testing results; mission impact deemed...insignificant due to multiple terminals at Shore site. Note for Sub (for MTBF, MTBCF and MTTR): Represents IOT &E hours; test duration limit for

  4. Downregulation of hTERT: an important As2O3 induced mechanism of apoptosis in myelodysplastic syndrome.

    Directory of Open Access Journals (Sweden)

    Weilai Xu

    Full Text Available Two myelodysplastic syndrome (MDS cell lines, MUTZ-1 and SKM-1 cells, were used to study the effect of arsenic trioxide (As2O3 on hematological malignant cells. As2O3 induced this two cell lines apoptosis via activation of caspase-3/8 and cleavage of poly (ADP-ribose polymerase (PARP, a DNA repair enzyme. As2O3 reduced NF-κB activity, which was important for inducing MUTZ-1 and SKM-1 cells apoptosis. As2O3 also inhibited the activities of hTERT in MUTZ-1 and SKM-1 cells. Moreover, the NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC, had no effect on caspase-8 activation, although PDTC did inhibit MUTZ-1 and SKM-1 cells proliferation. Incubation of MUTZ-1 cells with a caspase-8 inhibitor failed to block As2O3-induced inhibition of NF-κB activity. Our findings suggest that As2O3 may induce apoptosis in MUTZ-1 and SKM-1 cells by two independent pathways: first, by activation of caspase-3/8 and PARP; and second, by inhibition of NF-κB activity, which results in downregulation of hTERT expression. We conclude that hTERT and NF-κB are important molecular targets in As2O3-induced apoptosis.

  5. Kaempferol induces ATM/p53-mediated death receptor and mitochondrial apoptosis in human umbilical vein endothelial cells.

    Science.gov (United States)

    Lee, Chiu-Fang; Yang, Jai-Sing; Tsai, Fuu-Jen; Chiang, Ni-Na; Lu, Chi-Cheng; Huang, Yu-Syuan; Chen, Chun; Chen, Fu-An

    2016-05-01

    Kaempferol is a member of the flavonoid compounds found in vegetables and fruits. It is shown to exhibit biological impact and anticancer activity, but no report exists on the angiogenic effect of kaempferol and induction of cell apoptosis in vitro. In this study, we investigated the role of kaempferol on anti-angiogenic property and the apoptotic mechanism of human umbilical vein endothelial cells (HUVECs). Our results demonstrated that kaempferol decreased HUVEC viability in a time- and concentration-dependent manner. Kaempferol also induced morphological changes and sub-G1 phase cell population (apoptotic cells). Kaempferol triggered apoptosis of HUVECs as detecting by DNA fragmentation, comet assay and immunofluorescent staining for activated caspase-3. The caspase signals, including caspase-8, -9 and -3, were time-dependently activated in HUVECs after kaempferol exposure. Furthermore, pre-treatment with a specific inhibitor of caspase-8 (Z-IETD-FMK) significantly reduced the activity of caspase-8, -9 and -3, indicating that extrinsic pathway is a major signaling pathway in kaempferol-treated HUVECs. Importantly, kaempferol promoted reactive oxygen species (ROS) evaluated using flow cytometric assay in HUVECs. We further investigated the upstream extrinsic pathway and showed that kaempferol stimulated death receptor signals [Fas/CD95, death receptor 4 (DR4) and DR5] through increasing the levels of phosphorylated p53 and phosphorylated ATM pathways in HUVECs, which can be individually confirmed by N-acetylcysteine (NAC), ATM specific inhibitor (caffeine) and p53 siRNA. Based on these results, kaempferol-induced HUVEC apoptosis was involved in an ROS-mediated p53/ATM/death receptor signaling. Kaempferol might possess therapeutic effects on cancer treatment in anti-vascular targeting.

  6. Spy1 induces de-ubiquitinating of RIP1 arrest and confers glioblastoma's resistance to tumor necrosis factor (TNF-α)-induced apoptosis through suppressing the association of CLIPR-59 and CYLD

    Science.gov (United States)

    Ding, Zongmei; Liu, Yonghua; Yao, Li; Wang, Donglin; Zhang, Jianguo; Cui, Gang; Yang, Xiaojing; Huang, Xianting; Liu, Fang; Shen, Aiguo

    2015-01-01

    Glioblastoma multiforme (GBM), a grade-IV glioma, is resistant to TNF-α induced apoptosis. CLIPR-59 modulates ubiquitination of RIP1, thus promoting Caspase-8 activation to induce apoptosis by TNF-α. Here we reported that CLIPR-59 was down-regulated in GBM cells and high-grade glioma tumor samples, which was associated with decreased cancer-free survival. In GBM cells, CLIPR-59 interacts with Spy1, resulting in its decreased association with CYLD, a de-ubiquitinating enzyme. Moreover, experimental reduction of Spy1 levels decreased GBM cells viability, while increased the lysine-63-dependent de-ubiquitinating activity of RIP1 via enhancing the binding ability of CLIPR-59 and CYLD in GBM, thus promoting Caspase-8 and Caspase-3 activation to induce apoptosis by TNF-α. These findings have identified a novel Spy1-CLIPR-59 interplay in GBM cell's resistance to TNF-α-induced apoptosis revealing a potential target in the intervention of malignant brain tumors. PMID:26017671

  7. MORINGA TEA BLOCKS ACUTE LUNG INFLAMMATION INDUCED BY SWINE CONFINEMENT DUST THROUGH A MECHANISM INVOLVING TNF-α EXPRESSION, C-JUN N-TERMINAL KINASE ACTIVATION AND NEUTROPHIL REGULATION

    Directory of Open Access Journals (Sweden)

    Mykea Mcknight

    2014-01-01

    Full Text Available Plant based products represent a promising alternative to conventional treatments for inflammation. Moringa oleifera Lam is a tree rich in proteins, vitamins, minerals and a variety of phytochemcals with health benefits. Among the reported health benefits are antioxidant and anti-inflammatory properties. The purpose of this study was to investigate whether tea brewed from dried Moringa leaves would abrogate inflammation in a mouse model of acute lung inflammation induced by LPS or extracts prepared from dust collected from a swine confinement facility (DE. Mice were offered water or Moringa tea for seven days. Tea consumption was significantly greater than that of water consumption on days 1 and 6, but there were no significant differences in weight gain or food consumption. On day seven, mice from both groups were forced to inhale, via intranasal challenge, either Phosphate Buffered Saline (PBS, Lipopolysaccharide (LPS [10 µg mL-1] or DE [10%]. Compared to mice that drank water, mice that drank Moringa tea had significantly less protein (p<0.05 and cellular influx (p<0.0001 into the lung after inhalation of 10% DE. No difference in neutrophil migration into the lungs of water and M. tea groups after LPS or DE challenge was detected. But mice that drank tea had significantly (p<0.05 more neutrophils with apoptotic morphology after DE challenge. TNF-α expression 24 h after inhalation of 10% DE, was significantly higher (p<0.05 in lungs of M. tea mouse group as compared to water group. This increase in TNF-α was accompanied by higher levels of pro and anti-inflammatory cytokines. Finally, activation of c-Jun N-terminal Kinase (JNK in lungs of M. tea+DE group 24 h post inhalation was decreased. Taken together these results suggest that Moringa oleifera leaf tea exerts anti-inflammatory properties on acute lung inflammation induced by swine confinement dust through a mechanism involving neutrophil regulation and JNK

  8. Thymoquinone ameliorates testicular tissue inflammation induced by chronic administration of oral sodium nitrite.

    Science.gov (United States)

    Alyoussef, A; Al-Gayyar, M M H

    2016-06-01

    Although sodium nitrite has been widely used as food preservative, building bases of scientific evidence about nitrite continues to oppose the general safety in human health. Moreover, thymoquinone (TQ) has therapeutic potential as antioxidant, anti-inflammatory, antibacterial and anticancer. Therefore, we investigated the effects of both sodium nitrite and TQ on testicular tissues of rats. Forty adult male Sprague Dawley rats were used. They received either 80 mg kg(-1) sodium nitrite or 50 mg kg(-1) TQ daily for twelve weeks. Serum testosterone was measured. Testis were weighed and the testicular tissue homogenates were used for measurements of tumour necrosis factor (TNF)-α, interleukin (IL)-1β, IL-4, IL-6, IL10, caspase-3, caspase-8 and caspase-9. Sodium nitrite resulted in significant reduction in serum testosterone concentration and elevation in testis weight and Gonado-Somatic Index. We found significant reduction in testicular tissues levels of IL-4 and IL-10 associated with elevated levels of TNF-α, IL-1β, IL-6, caspase-3, caspase-8 and caspase-9. In conclusion, chronic oral sodium nitrite induced changes in the weight of rat testis accompanied by elevation in the testicular tissue level of oxidative stress markers and inflammatory cytokines. TQ attenuated sodium nitrite-induced testicular tissue damage through blocking oxidative stress, restoration of normal inflammatory cytokines balance and blocking of apoptosis.

  9. XIAP acts as a switch between type I and type II FAS-induced apoptosis signalling

    Science.gov (United States)

    Jost, Philipp J.; Grabow, Stephanie; Gray, Daniel; McKenzie, Mark D.; Nachbur, Ueli; Huang, David C.S.; Bouillet, Philippe; Thomas, Helen E.; Borner, Christoph; Silke, John; Strasser, Andreas; Kaufmann, Thomas

    2010-01-01

    FAS (APO-1/CD95) and its physiological ligand, FASL, regulate apoptotic death of unwanted or dangerous cells in many tissues, functioning as a guardian against autoimmunity and cancer development1-4. Distinct cell types differ in the mechanisms by which the ‘death receptor’ FAS triggers their apoptosis1-4. In type I cells, such as lymphocytes, activation of ‘effector caspases’ by FAS-induced activation of caspase-8 suffices for cell killing whereas in type II cells, including hepatocytes and pancreatic β-cells, amplification of the caspase cascade through caspase-8 mediated activation of the pro-apoptotic BCL-2 family member BID5 is essential6-8. Here we show, that loss of X-chromosome linked inhibitor of apoptosis (XIAP)9,10 function by gene-targeting or treatment with a second mitochondria-derived activator of caspases (SMAC11, also called DIABLO12: direct IAP binding protein with low pI) mimetic drug rendered hepatocytes independent of BID for FAS-induced apoptosis signalling. These results show that XIAP is the critical discriminator between type I versus type II apoptosis signalling and suggest that IAP inhibitors should be used with caution in cancer patients with underlying liver conditions. PMID:19626005

  10. XIAP discriminates between type I and type II FAS-induced apoptosis.

    Science.gov (United States)

    Jost, Philipp J; Grabow, Stephanie; Gray, Daniel; McKenzie, Mark D; Nachbur, Ueli; Huang, David C S; Bouillet, Philippe; Thomas, Helen E; Borner, Christoph; Silke, John; Strasser, Andreas; Kaufmann, Thomas

    2009-08-20

    FAS (also called APO-1 and CD95) and its physiological ligand, FASL, regulate apoptosis of unwanted or dangerous cells, functioning as a guardian against autoimmunity and cancer development. Distinct cell types differ in the mechanisms by which the 'death receptor' FAS triggers their apoptosis. In type I cells, such as lymphocytes, activation of 'effector caspases' by FAS-induced activation of caspase-8 suffices for cell killing, whereas in type II cells, including hepatocytes and pancreatic beta-cells, caspase cascade amplification through caspase-8-mediated activation of the pro-apoptotic BCL-2 family member BID (BH3 interacting domain death agonist) is essential. Here we show that loss of XIAP (X-chromosome linked inhibitor of apoptosis protein) function by gene targeting or treatment with a second mitochondria-derived activator of caspases (SMAC, also called DIABLO; direct IAP-binding protein with low pI) mimetic drug in mice rendered hepatocytes and beta-cells independent of BID for FAS-induced apoptosis. These results show that XIAP is the critical discriminator between type I and type II apoptosis signalling and suggest that IAP inhibitors should be used with caution in cancer patients with underlying liver conditions.

  11. Novel targets for endoplasmic reticulum stress-induced apoptosis in B-CLL.

    Science.gov (United States)

    Rosati, Emanuela; Sabatini, Rita; Rampino, Giuliana; De Falco, Filomena; Di Ianni, Mauro; Falzetti, Franca; Fettucciari, Katia; Bartoli, Andrea; Screpanti, Isabella; Marconi, Pierfrancesco

    2010-10-14

    A better understanding of apoptotic signaling in B-chronic lymphocytic leukemia (B-CLL) cells may help to define new therapeutic strategies. This study investigated endoplasmic reticulum (ER) stress signaling in spontaneous apoptosis of B-CLL cells and whether manipulating ER stress increases their apoptosis. Results show that a novel ER stress-triggered caspase cascade, initiated by caspase-4 and involving caspase-8 and -3, plays an important role in spontaneous B-CLL cell apoptosis. ER stress-induced apoptosis in B-CLL cells also involves CHOP/GADD153 up-regulation, increased JNK1/2 phosphorylation, and caspase-8-mediated cleavage of Bap31 to Bap20, known to propagate apoptotic signals from ER to mitochondria. In ex vivo B-CLL cells, some apoptotic events associated with mitochondrial pathway also occur, including mitochondrial cytochrome c release and caspase-9 processing. However, pharmacologic inhibition studies show that caspase-9 plays a minor role in B-CLL cell apoptosis. ER stress also triggers survival signals in B-CLL cells by increasing BiP/GRP78 expression. Manipulating ER signaling by siRNA down-regulation of BiP/GRP78 or treating B-CLL cells with 2 well-known ER stress-inducers, tunicamycin and thapsigargin, increases their apoptosis. Overall, our findings show that ER triggers an essential pathway for B-CLL cell apoptosis and suggest that genetic and pharmacologic manipulation of ER signaling could represent an important therapeutic strategy.

  12. Copper deficiency induced emphysema is associated with focal adhesion kinase inactivation.

    Directory of Open Access Journals (Sweden)

    Shiro Mizuno

    Full Text Available BACKGROUND: Copper is an important regulator of hypoxia inducible factor 1 alpha (HIF-1α dependent vascular endothelial growth factor (VEGF expression, and is also required for the activity of lysyl oxidase (LOX to effect matrix protein cross-linking. Cell detachment from the extracellular matrix can induce apoptosis (anoikis via inactivation of focal adhesion kinase (FAK. METHODOLOGY: To examine the molecular mechanisms whereby copper depletion causes the destruction of the normal alveolar architecture via anoikis, Male Sprague-Dawley rats were fed a copper deficient diet for 6 weeks while being treated with the copper chelator, tetrathiomolybdate. Other groups of rats were treated with the inhibitor of auto-phosphorylation of FAK, 1,2,4,5-benzenetetraamine tetrahydrochloride (1,2,4,5-BT or FAK small interfering RNA (siRNA. PRINCIPAL FINDINGS: Copper depletion caused emphysematous changes, decreased HIF-1α activity, and downregulated VEGF expression in the rat lungs. Cleaved caspase-3, caspase-8 and Bcl-2 interacting mediator of cell death (Bim expression was increased, and the phosphorylation of FAK was decreased in copper depleted rat lungs. Administration of 1,2,4,5-BT and FAK siRNA caused emphysematous lung destruction associated with increased expression of cleaved capase-3, caspase-8 and Bim. CONCLUSIONS: These data indicate that copper-dependent mechanisms contribute to the pathogenesis of emphysema, which may be associated with decreased HIF-1α and FAK activity in the lung.

  13. 探讨无痛人工流产用于终止早期妊娠的临床应用价值%The Clinical Value of the Induced Abortion in the Termination Early Pregnancy

    Institute of Scientific and Technical Information of China (English)

    胡蓉

    2013-01-01

    Objective:To observe the application value of the induced abortion used to the early pregnancy for clinical make guidance.Method:One hundred and four healthy pregnant women admitted to our hospital from March 2010 to March 2013 were selected,they were divided into the abortion group and the medical abortion group,52 cases in each group.The abortion group was given induced abortion,the medical abortion group was given Mifepristone Tablets and Misoprostol Tablets,The indexes(the amount of vaginal bleeding,vaginal bleeding time,menstrual recovery time,duration of abdominal pain),the rate of complete abortion and the incidence of adverse reactions were compared between the two groups after treatment.Result:The abortion group was significantly better than the medical abortion group after treatment,the difference was statistically significant(P<0.05);The abortion rate of the abortion group was 98.08%,it was significantly higher than the medical abortion group(80.77%),the difference was statistically significant (P<0.05);the adverse reaction rate of the abortion group was 11.54%,it was significantly lower than the medical abortion group(36.54%),the difference was statistically significant(P<0.05).Conclusion:Painless artificial abortion for termination of early pregnancy clinical effect significantly, has the characteristics of simple operation,less bleeding,complete abortion rate is high,less adverse reaction,and it is worthy of wide application.%目的:分析总结无痛人工流产用于终止早期妊娠的临床应用价值,为临床推广做出指导。方法:选取本站2010年3月-2013年3月收治的104例自愿终止妊娠的健康孕妇,按照随机数字表法将其分为人工流产组和药物流产组各52例,人工流产组给予无痛人工流产,药物流产组给予米非司酮联合米索前列醇口服,观察比较两组患者治疗后各项指标(阴道出血量、阴道出血时间、月经恢复时间、腹痛持续时间)、治

  14. Is Lake Tahoe Terminal?

    Science.gov (United States)

    Coats, R. N.; Reuter, J.; Heyvaert, A.; Lewis, J.; Sahoo, G. B.; Schladow, G.; Thorne, J. H.

    2014-12-01

    ) the climatic water deficit will increase, especially at high elevations that will be most affected by the loss of snow, with likely consequences for existing vegetation and fire frequency. Hydrologically, Lake Tahoe is intermittently terminal; in a medical sense it is not yet terminal, but its condition—especially its valued clarity and deep blue color--is serious.

  15. Autophagy protein p62/SQSTM1 is involved in HAMLET-induced cell death by modulating apotosis in U87MG cells.

    Science.gov (United States)

    Zhang, Y-B; Gong, J-L; Xing, T-Y; Zheng, S-P; Ding, W

    2013-03-21

    HAMLET is a complex of oleic acids and decalcified α-lactalbumin that was discovered to selectively kill tumor cells both in vitro and in vivo. Autophagy is an important cellular process involved in drug-induced cell death of glioma cells. We treated U87MG human glioma cells with HAMLET and found that the cell viability was significantly decreased and accompanied with the activation of autophagy. Interestingly, we observed an increase in p62/SQSTM1, an important substrate of autophagosome enzymes, at the protein level upon HAMLET treatment for short periods. To better understand the functionality of autophagy and p62/SQSTM1 in HAMLET-induced cell death, we modulated the level of autophagy or p62/SQSTM1 with biochemical or genetic methods. The results showed that inhibition of autophagy aggravated HAMLET-induced cell death, whereas activation of authophagy attenuated this process. Meanwhile, we found that overexpression of wild-type p62/SQSTM1 was able to activate caspase-8, and then promote HAMLET-induced apoptosis, whereas knockdown of p62/SQSTM1 manifested the opposite effect. We further demonstrated that the function of p62/SQSTM1 following HAMLET treatment required its C-terminus UBA domain. Our results indicated that in addition to being a marker of autophagy activation in HAMLET-treated glioma cells, p62/SQSTM1 could also function as an important mediator for the activation of caspase-8-dependent cell death.

  16. Oral Lactobacillus reuteri GMN-32 treatment reduces blood glucose concentrations and promotes cardiac function in rats with streptozotocin-induced diabetes mellitus.

    Science.gov (United States)

    Lin, Chih-Hsueh; Lin, Cheng-Chieh; Shibu, Marthandam Asokan; Liu, Chiu-Shong; Kuo, Chia-Hua; Tsai, Fuu-Jen; Tsai, Chang-Hai; Hsieh, Cheng-Hong; Chen, Yi-Hsing; Huang, Chih-Yang

    2014-02-01

    Impaired regulation of blood glucose levels in diabetes mellitus (DM) patients and the associated elevation of blood glucose levels are known to increase the risk of diabetic cardiomyopathy (DC). In the present study, a probiotic bacterium, Lactobacillus reuteri GMN-32, was evaluated for its potential to reduce blood glucose levels and to provide protection against DC risks in streptozotocin (STZ)-induced DM rats. The blood glucose levels of the STZ-induced DM rats when treated with L. reuteri GMN-32 decreased from 4480 to 3620 mg/l (with 10⁷ colony-forming units (cfu)/d) and 3040 mg/l (with 10⁹ cfu/d). Probiotic treatment also reduced the changes in the heart caused by the effects of DM. Furthermore, the Fas/Fas-associated protein with death domain pathway-induced caspase 8-mediated apoptosis that was observed in the cardiomyocytes of the STZ-induced DM rats was also found to be controlled in the probiotic-treated rats. The results highlight that L. reuteri GMN-32 treatment reduces blood glucose levels, inhibits caspase 8-mediated apoptosis and promotes cardiac function in DM rats as observed from their ejection fraction and fractional shortening values. In conclusion, the administration of L. reuteri GMN-32 probiotics can regulate blood glucose levels, protect cardiomyocytes and prevent DC in DM rats.

  17. Terminal Satisfiability in GSTE

    Directory of Open Access Journals (Sweden)

    Yongsheng Xu

    2014-01-01

    Full Text Available Generalized symbolic trajectory evaluation (GSTE is an extension of symbolic trajectory evaluation (STE and a method of model checking. GSTE specifications are given as assertion graphs. There are four efficient methods to verify whether a circuit model obeys an assertion graph in GSTE, Model Checking Strong Satisfiability (SMC, Model Checking Normal Satisfiability (NMC, Model Checking Fair Satisfiability (FMC, and Model Checking Terminal Satisfiability (TMC. SMC, NMC, and FMC have been proved and applied in industry, but TMC has not. This paper gives a six-tuple definition and presents a new algorithm for TMC. Based on these, we prove that our algorithm is sound and complete. It solves the SMC’s limitation (resulting in false negative without extending from finite specification to infinite specification. At last, a case of using TMC to verify a realistic hardware circuit round-robin arbiter is achieved. Avoiding verifying the undesired paths which are not related to the specifications, TMC makes it possible to reduce the computational complexity, and the experimental results suggest that the time cost by SMC is 3.14× with TMC in the case.

  18. War Termination: A Selected Bibliography

    Science.gov (United States)

    2010-10-01

    Peace Research 33, no. 4 (November 1996): 491-496. JSTOR Phinney, Catherine. "Enhancing Conflict Termination through Problem Solving...96." Journal of Peace Research 34, no. 3 (August 1997): 339-358. JSTOR Bibliographies Bibliography Branch, comp. Conflict Termination

  19. Selection of Air Terminal Device

    DEFF Research Database (Denmark)

    Nielsen, Peter V.

    This paper discusses the selection of the air terminal device for the experiments and numerical prediction in the International Energy Agency Annex 20 work: Air Flow Pattern within Buildings,......This paper discusses the selection of the air terminal device for the experiments and numerical prediction in the International Energy Agency Annex 20 work: Air Flow Pattern within Buildings,...

  20. Overexpression of cellular repressor of E1A-stimulated genes inhibits TNF-{alpha}-induced apoptosis via NF-{kappa}B in mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Cheng-Fei [Xijing Hospital, Fourth Military Medical University, Xi' an (China); Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China); Han, Ya-Ling, E-mail: hanyaling53@gmail.com [Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China); Jie-Deng,; Yan, Cheng-Hui; Jian-Kang,; Bo-Luan,; Jie-Li [Cardiovascular Research Institute and Department of Cardiology, Shenyang Northern Hospital, Shenyang (China)

    2011-03-25

    Research highlights: {yields} CREG protected MSCs from tumor necrosis factor-{alpha} (TNF-{alpha}) induced apoptosis. {yields} CREG inhibits the phosphorylation of I{kappa}B{alpha} and prevents the activation of NF-{kappa}B. {yields} CREG inhibits NF-{kappa}B nuclear translocation and pro-apoptosis protein transcription. {yields} CREG anti-apoptotic effect involves inhibition of the death receptor pathway. {yields} p53 is downregulated by CREG via NF-{kappa}B pathway under TNF-{alpha} stimulation. -- Abstract: Bone marrow-derived mesenchymal stem cells (MSCs) show great potential for therapeutic repair after myocardial infarction. However, poor viability of transplanted MSCs in the ischemic heart has limited their use. Cellular repressor of E1A-stimulated genes (CREG) has been identified as a potent inhibitor of apoptosis. This study therefore aimed to determine if rat bone marrow MSCs transfected with CREG-were able to effectively resist apoptosis induced by inflammatory mediators, and to demonstrate the mechanism of CREG action. Apoptosis was determined by flow cytometric and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling assays. The pathways mediating these apoptotic effects were investigated by Western blotting. Overexpression of CREG markedly protected MSCs from tumor necrosis factor-{alpha} (TNF-{alpha}) induced apoptosis by 50% after 10 h, through inhibition of the death-receptor-mediated apoptotic pathway, leading to attenuation of caspase-8 and caspase-3. Moreover, CREG resisted the serine phosphorylation of I{kappa}B{alpha} and prevented the nuclear translocation of the transcription factor nuclear factor-{kappa}B (NF-{kappa}B) under TNF-{alpha} stimulation. Treatment of cells with the NF-{kappa}B inhibitor pyrrolidine dithiocarbamate (PDTC) significantly increased the transcription of pro-apoptosis proteins (p53 and Fas) by NF-{kappa}B, and attenuated the anti-apoptotic effects of CREG on MSCs. The results of this study

  1. Conformal Mapping for Multiple Terminals

    CERN Document Server

    Wang, Weimin; Wang, Qiang; Ren, Hao

    2015-01-01

    Conformal mapping is an important mathematical tool in many physical and engineering fields, especially in electrostatics, fluid mechanics, classical mechanics, and transformation optics. However in the existing textbooks and literatures, it is only adopted to solve the problems which have only two terminals. Two terminals with electric potential differences, pressure difference, optical path difference, etc., can be mapped conformally onto a solvable structure, e.g., a rectangle, where the two terminals are mapped onto two opposite edges of the rectangle. Here we show a conformal mapping method for multiple terminals, which is more common in practical applications. Through accurate analysis of the boundary conditions, additional terminals or boundaries are folded in the inner of the mapped rectangle. Then the solution will not be influenced. The method is described in several typical situations and two application examples are detailed. The first example is an electrostatic actuator with three electrodes. A ...

  2. Fluorescence Resonance Energy Transfer Analysis of Bid Activation in Living Cells during Ultraviolet-induced Apoptosis

    Institute of Scientific and Technical Information of China (English)

    Yinyuan WU; Da XING; Lei LIU; Tongsheng CHEN; Wei R. CHEN

    2007-01-01

    Ultraviolet (UV) irradiation is a DNA-damaging agent that triggers apoptosis through both the membrane death receptor and mitochondrial apoptotic signaling pathways. Bid, a pro-apoptotic Bcl-2family member, is important in most cell types to apoptosis in response to DNA damage. In this study, a recombinant plasmid, YFP-Bid-CFP, comprised of yellow and cyan fluorescent protein and a full length Bid,was used as a fluorescence resonance energy transfer analysis (FRET) probe. Using the FRET technique based on YFP-Bid-CFP, we found that Bid activation was initiated at 9±1 h after UV irradiation, and the average duration of the activation was 75± 10 min. Bid activation coincided with a collapse of the mitochondrial membrane potential with an average duration of 50±10 min. When cells were pretreated with Z-IETD-fmk(caspase-8 specific inhibitor) the process of Bid activation was completely inhibited, but the apoptosis was only partially affected. Z-DEVD-fmk (caspase-3 inhibitor) and Z-FA-fmk (non asp specific inhibitor) did not block Bid activation. Furthermore, the endogenous Bid activation with or without Z-IETD-fmk in response to UV irradiation was confirmed by Western blotting. In summary, using the FRET technique, we observed the dynamics of Bid activation during UV-induced apoptosis and found that it was a caspase-8 dependent event.

  3. The venom of the spider Macrothele raveni induces apoptosis in the myelogenous leukemia K562 cell line.

    Science.gov (United States)

    Liu, Zhonghua; Zhao, Yan; Li, Jing; Xu, Shiyan; Liu, Changjun; Zhu, Yanghui; Liang, Songping

    2012-08-01

    Spider venoms are a rich source of bioactive compounds with therapeutic potential. In traditional Chinese medicine, spiders and spider venoms have been used in the treatment of various ailments. In the present study, the venom of the spider Macrothele raveni potently suppressed cell growth in the myelogenous leukemia K562 cell line in a dose and time-dependent manner with an IC(50) of 5.1 μg/mL. The venom also had a low inhibitory effect on human lymphocytes with an IC(50) of approximately 36.4 μg/mL, indicating that the venom is relatively selective for leukemic cells. Venom treated K562 cells showed typical morphological indicators of apoptosis including condensation of nuclei and fragmentation of DNA. Annexin V-FITC and propidium iodide dual staining further demonstrated that the venom had potent apoptogenic activity. Venom treatment induced caspase 3 and caspase 8 activation in K562 cells and promoted PARP cleavage. The present results indicate that the venom of the spider M. raveni potently and selectively suppresses the growth of K562 cells by inducing apoptosis via caspase 3 and caspase 8 mediated signaling pathways.

  4. α-Hispanolol sensitizes hepatocellular carcinoma cells to TRAIL-induced apoptosis via death receptor up-regulation

    Energy Technology Data Exchange (ETDEWEB)

    Mota, Alba, E-mail: amota@iib.uam.es [Unidad de Terapias Farmacológicas, Área de Genética Humana, Instituto de Investigación de Enfermedades Raras (IIER), Instituto de Salud Carlos III, Madrid (Spain); Jiménez-Garcia, Lidia, E-mail: ljimenez@isciii.es [Unidad de Terapias Farmacológicas, Área de Genética Humana, Instituto de Investigación de Enfermedades Raras (IIER), Instituto de Salud Carlos III, Madrid (Spain); Herránz, Sandra, E-mail: sherranz@isciii.es [Unidad de Terapias Farmacológicas, Área de Genética Humana, Instituto de Investigación de Enfermedades Raras (IIER), Instituto de Salud Carlos III, Madrid (Spain); Heras, Beatriz de las, E-mail: lasheras@ucm.es [Departamento de Farmacología, Facultad de Farmacia, Universidad Complutense de Madrid (UCM), Madrid (Spain); Hortelano, Sonsoles, E-mail: shortelano@isciii.es [Unidad de Terapias Farmacológicas, Área de Genética Humana, Instituto de Investigación de Enfermedades Raras (IIER), Instituto de Salud Carlos III, Madrid (Spain)

    2015-08-01

    Hispanolone derivatives have been previously described as anti-inflammatory and antitumoral agents. However, their effects on overcoming Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) resistance remain to be elucidated. In this study, we analyzed the cytotoxic effects of the synthetic hispanolone derivative α-hispanolol (α-H) in several tumor cell lines, and we evaluated the induction of apoptosis, as well as the TRAIL-sensitizing potential of α-H in the hepatocellular carcinoma cell line HepG2. Our data show that α-H decreased cell viability in a dose-dependent manner in HeLa, MDA-MB231, U87 and HepG2 cell lines, with a more prominent effect in HepG2 cells. Interestingly, α-H had no effect on non-tumoral cells. α-H induced activation of caspase-8 and caspase-9 and also increased levels of the proapoptotic protein Bax, decreasing antiapoptotic proteins (Bcl-2, X-IAP and IAP-1) in HepG2 cells. Specific inhibition of caspase-8 abrogated the cascade of caspase activation, suggesting that the extrinsic pathway has a critical role in the apoptotic events induced by α-H. Furthermore, combined treatment of α-H with TRAIL enhanced apoptosis in HepG2 cells, activating caspase-8 and caspase-9. This correlated with up-regulation of both the TRAIL death receptor DR4 and DR5. DR4 or DR5 neutralizing antibodies abolished the effect of α-H on TRAIL-induced apoptosis, suggesting that sensitization was mediated through the death receptor pathway. Our results demonstrate that α-H induced apoptosis in the human hepatocellular carcinoma cell line HepG2 through activation of caspases and induction of the death receptor pathway. In addition, we describe a novel function of α-H as a sensitizer on TRAIL-induced apoptotic cell death in HepG2 cells. - Highlights: • α-Hispanolol induced apoptosis in the human hepatocellular carcinoma cell line HepG2. • α-Hispanolol induced activation of caspases and the death receptor pathway. • α-Hispanolol enhanced

  5. Antiapoptotic mechanism of cannabinoid receptor 2 agonist on cisplatin-induced apoptosis in the HEI-OC1 auditory cell line.

    Science.gov (United States)

    Jeong, Hyun-Ja; Kim, Su-Jin; Moon, Phil-Dong; Kim, Na-Hyun; Kim, Jung-Sun; Park, Rae-Kil; Kim, Min-Sun; Park, Byung-Rim; Jeong, Sejin; Um, Jae-Young; Kim, Hyung-Min; Hong, Seung-Heon

    2007-03-01

    Cisplatin is a highly effective chemotherapeutic agent but with significant ototoxic side effects. Apoptosis is an important mechanism of cochlear hair cell loss following exposure to an ototoxic level of cisplatin. The present study investigated the effects of the cannabinoid receptor 2 (CB2) ligand JWH-015 on cisplatin-induced apoptosis. CB2 mRNA was constitutively expressed in the auditory cell line HEI-OC1. By using MTT assay, DNA fragmentation, and FACS analysis, we demonstrated that apoptosis induced by cisplatin was inhibited by treatment with JWH-015 in a dose-dependent manner. Activation of caspase-3, caspase-8, and caspase-9 was detected after treatment with cisplatin, and the cleavage of poly-(ADP)-ribose polymerase (PARP) was observed within cisplatin-treated HEI-OC1 cells. JWH-015 inhibited the activation of caspase-3, caspase-8, and caspase-9; cleavage of PARP; and release of cytochrome c. JWH-015 also inhibited the apoptosis through activation of the extracellular signal-regulated kinase pathway. Finally, JWH-015 inhibited cisplatin-induced reactive oxygen species and tumor necrosis factor-alpha production. Collectively, these findings show that blocking a critical step in apoptosis by using JWH-015 may be a useful strategy to prevent harmful side effects of cisplatin ototoxicity in patients having to undergo chemotherapy.

  6. Phosphorylated extracellular signal-regulated kinase up-regulated p53 expression in shikonin-induced HeLa cell apoptosis

    Institute of Scientific and Technical Information of China (English)

    WU Zhen; WU Li-jun; TASHIRO Shinichi; ONODERA Satoshi; IKEJIMA Takashi

    2005-01-01

    Background The role of extracellular signal-regulated kinase 1/2 (ERK1/2) in shikonin-induced HeLa cells apoptosis remains vague. This study was to investigate the activation of caspase pathways and the role of ERK1/2 in human cervical cancer cells, HeLa, by shikonin.Methods The inhibitory effect of shikonin on the growth of HeLa cells was measured by MTT assay. Fluorescent microscopic analysis of apoptotic cells stained with 4’,6’-oliiamiclino-2-phenylindole C (DAPI) and Hoechst 33258 was carried out. Caspase-3 and -8 activities were detected using caspase-3 substrate and caspase-8 substrate as substrates, respectively. The protein levels of ERK, p53 and p-ERK were determined by Western blot analysis.Results Shikonin inhibited cell growth in a time- and dose-dependent manner. Caspase-3 and caspase-8 were activated in the apoptotic process and caspase inhibitors effectively reversed shikonin-induced apoptosis. Phosphorylation of ERK resulted in up-regulation of p53 expression, which was blocked by mitogen-activated protein kinase (MEK), inhibitor PD 98059.Conclusion Shikonin induces HeLa cell apoptosis through the ERK, p53 and caspase pathways.

  7. The use of mobile satellite communication terminals

    Science.gov (United States)

    Law, P. A.

    The role of small portable terminals in military satellite systems is examined; the discussion embraces terminals with an antenna reflector diameter of seven meters or less. Emphasis is placed on the specification of MARMOSET (Marconi Mobile Satellite Earth Terminal). Also considered are ship-borne satellite terminals, the improved SCOT terminal, interoperability, reduced downlink power, and reliability and availability.

  8. Low Earth Orbiter: Terminal

    Science.gov (United States)

    Kremer, Steven E.; Bundick, Steven N.

    1999-01-01

    In response to the current government budgetary environment that requires the National Aeronautics and Space Administration (NASA) to do more with less, NASA/Goddard Space Flight Center's Wallops Flight Facility has developed and implemented a class of ground stations known as a Low Earth Orbiter-Terminal (LEO-T). This development thus provides a low-cost autonomous ground tracking service for NASA's customers. More importantly, this accomplishment provides a commercial source to spacecraft customers around the world to purchase directly from the company awarded the NASA contract to build these systems. A few years ago, NASA was driven to provide more ground station capacity for spacecraft telemetry, tracking, and command (TT&C) services with a decreasing budget. NASA also made a decision to develop many smaller, cheaper satellites rather than a few large spacecraft as done in the past. In addition, university class missions were being driven to provide their own TT&C services due to the increasing load on the NASA ground-tracking network. NASA's solution for this ever increasing load was to use the existing large aperture systems to support those missions requiring that level of performance and to support the remainder of the missions with the autonomous LEO-T systems. The LEO-T antenna system is a smaller, cheaper, and fully autonomous unstaffed system that can operate without the existing NASA support infrastructure. The LEO-T provides a low-cost, reliable space communications service to the expanding number of low-earth orbiting missions around the world. The system is also fostering developments that improve cost-effectiveness of autonomous-class capabilities for NASA and commercial space use. NASA has installed three LEO-T systems. One station is at the University of Puerto Rico, the second system is installed at the Poker Flat Research Range near Fairbanks, Alaska, and the third system is installed at NASA's Wallops Flight Facility in Virginia. This paper

  9. Amino-terminated diamond surfaces: Photoelectron emission and photocatalytic properties

    Science.gov (United States)

    Zhu, Di; Bandy, Jason A.; Li, Shuo; Hamers, Robert J.

    2016-08-01

    We report a new approach to making stable negative electron-affinity diamond surfaces by terminating diamond with amino groups (also known as amine groups, -NH2). Previous studies have shown that negative electron affinity can be induced by terminating diamond surfaces with hydrogen, creating a surface dipole favorable toward electron emission. Here, we demonstrate that covalent tethering of positive charges in the form of protonated amino groups, -NH3+, also leads to negative electron affinity (NEA) and facile electron emission into vacuum and into water. Amino-terminated diamond was prepared using a very mild plasma discharge. Valence-band photoemission studies of the amino-terminated diamond samples show a characteristic "NEA" peak, demonstrating that the amino-terminated surface has NEA. Diamond's ability to emit electrons into water was evaluated using photochemical conversion of N2 to NH3. Time-resolved surface photovoltage studies were used to characterize charge separation at the diamond interface, and Mott-Schottky measurements were performed to characterize band-bending at the diamond-water interface. XPS studies show that the amino-terminated surfaces provide increased chemical resistance to oxidation compared with H-terminated diamond when illuminated with ultraviolet light.

  10. Bid and calpains cooperate to trigger oxaliplatin-induced apoptosis of cervical carcinoma HeLa cells.

    Science.gov (United States)

    Anguissola, Sergio; Köhler, Barbara; O'Byrne, Robert; Düssmann, Heiko; Cannon, Mary D; Murray, Frank E; Concannon, Caoimhin G; Rehm, Markus; Kögel, Donat; Prehn, Jochen H M

    2009-11-01

    The Bcl-2 homology 3-only protein Bid is an important mediator of death receptor-induced apoptosis. Recent reports and this study suggest that Bid may also mediate genotoxic drug-induced apoptosis of various human cancer cells. Here, we characterized the role of Bid and the mechanism of Bid activation during oxaliplatin-induced apoptosis of HeLa cervical cancer cells. Small hairpin RNA-mediated silencing of Bid protected HeLa cells against both death receptor- and oxaliplatin-induced apoptosis. Expression of a Bid mutant in which caspase-8 cleavage site was mutated (D59A) reactivated oxaliplatin-induced apoptosis in Bid-deficient cells but failed to reactivate death receptor-induced apoptosis, suggesting that caspase-8-mediated Bid cleavage did not contribute to oxaliplatin-induced apoptosis. Overexpression of bcl-2 or treatment with the pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-dl-Asp-fluoromethylketone abolished caspase-2, -8, -9, and -3 activation as well as Bid cleavage in response to oxaliplatin, suggesting that Bid cleavage occurred downstream of mitochondrial permeabilization and was predominantly mediated by caspases. We also detected an early activation of calpains in response to oxaliplatin. Calpain inhibition reduced Bid cleavage, mitochondrial depolarization, and activation of caspase-9, -3, -2, and -8 in response to oxaliplatin. Further experiments, however, suggested that Bid cleavage by calpains was not a prerequisite for oxaliplatin-induced apoptosis: single-cell imaging experiments using a yellow fluorescent protein-Bid-cyan fluorescent protein probe demonstrated translocation of full-length Bid to mitochondria that was insensitive to calpain or caspase inhibition. Moreover, calpain inhibition showed a potent protective effect in Bid-silenced cells. In conclusion, our data suggest that calpains and Bid act in a cooperative, but mutually independent, manner to mediate oxaliplatin-induced apoptosis of HeLa cells.

  11. [Mechanism of HL-60 cells apoptosis induced by proteasome inhibitor MG132].

    Science.gov (United States)

    Zhou, Yong-Ming; Yu, Mei-Xia; Qiu, Yu-Zhen; Xing, Xiao-Lei; Yao, Chun-Hong; Bai, Ru-Jun

    2013-08-01

    The purpose of this study was to elucidate the apoptosis, apoptotic pathway of HL-60 cells induced by proteasome inhibitor MG132 and its effect on allogeneic mixed lymphocyte reaction. Apoptosis of HL-60 cells was detected by flow cytometry, the expression of P21, P27 and P53 proteins in HL-60 cells treated with MG132 was assayed by Western blot. The HL-60 cells were treated with 1 µmol/L MG132 for 48 h, and irradiated by 75 Gy of (60)Co γ-ray, but their antigenicity was preserved. The effect of irradiated HL-60 cells treated with MG132 on proliferation of peripheral blood mononuclear cells (PBMNC) was measured by CCK-8 method. The results showed that the apoptotic rate of MG132-treated HL-60 cells increased in dose-and time-dependent manner. No significant changes in MG132-induced apoptosis were observed after inhibiting caspase-8 and caspase-9 pathway. The expression of P21 and P27 protein increased after treatment of HL-60 cells with MG132. CCK-8 test showed that HL-60 cells induced with low-dose of MG132 displayed the enhancing effect on proliferation of PBMNC. It is concluded that high dose of MG132 can induce the apoptosis of HL-60 cells, and has direct killing effect on HL-60 cells, but this inducing apoptotic effect on HL-60 cells can not be realized through caspase-8 and caspase-9 pathway. The P21 and P27 protein may be involved in MG132 induced HL-60 cell apoptosis. Low dose of MG132 promotes the proliferation of PBMNC in healthy individuals and enhance the immunity of organism.

  12. 75 FR 13644 - TORP Terminal LP, Bienville Offshore Energy Terminal Liquefied Natural Gas Deepwater Port License...

    Science.gov (United States)

    2010-03-22

    ... Maritime Administration TORP Terminal LP, Bienville Offshore Energy Terminal Liquefied Natural Gas...) for the TORP Terminal LP, Bienville Offshore Energy Terminal (BOET) Liquefied Natural Gas (LNG... Natural Gas Pipeline, Williams Natural Gas Pipeline, Destin Natural Gas Pipeline, and Viosca...

  13. Balanced Branching in Transcription Termination

    CERN Document Server

    Harrington, K J; Liang, S

    2000-01-01

    The theory of stochastic transcription termination based on free-energy competition requires two or more reaction rates to be delicately balanced over a wide range of physical conditions. A large body of work on glasses and large molecules suggests that this should be impossible in such a large system in the absence of a new organizing principle of matter. We review the experimental literature of termination and find no evidence for such a principle but many troubling inconsistencies, most notably anomalous memory effects. These suggest that termination has a deterministic component and may conceivably be not stochastic at all. We find that a key experiment by Wilson and von Hippel allegedly refuting deterministic termination was an incorrectly analyzed regulatory effect of Mg2+ binding.

  14. The natural triterpene maslinic acid induces apoptosis in HT29 colon cancer cells by a JNK-p53-dependent mechanism

    Directory of Open Access Journals (Sweden)

    Cascante Marta

    2011-04-01

    Full Text Available Abstract Background Maslinic acid, a pentacyclic triterpene found in the protective wax-like coating of the leaves and fruit of Olea europaea L., is a promising agent for the prevention of colon cancer. We have shown elsewhere that maslinic acid inhibits cell proliferation to a significant extent and activates mitochondrial apoptosis in colon cancer cells. In our latest work we have investigated further this compound's apoptotic molecular mechanism. Methods We used HT29 adenocarcinoma cells. Changes genotoxicity were analyzed by single-cell gel electrophoresis (comet assay. The cell cycle was determined by flow cytometry. Finally, changes in protein expression were examined by western blotting. Student's t-test was used for statistical comparison. Results HT29 cells treated with maslinic acid showed significant increases in genotoxicity and cell-cycle arrest during the G0/G1 phase after 72 hours' treatment and an apoptotic sub-G0/G1 peak after 96 hours. Nevertheless, the molecular mechanism for this cytotoxic effect of maslinic acid has never been properly explored. We show here that the anti-tumoral activity of maslinic acid might proceed via p53-mediated apoptosis by acting upon the main signaling components that lead to an increase in p53 activity and the induction of the rest of the factors that participate in the apoptotic pathway. We found that in HT29 cells maslinic acid activated the expression of c-Jun NH2-terminal kinase (JNK, thus inducing p53. Treatment of tumor cells with maslinic acid also resulted in an increase in the expression of Bid and Bax, repression of Bcl-2, release of cytochrome-c and an increase in the expression of caspases -9, -3, and -7. Moreover, maslinic acid produced belated caspase-8 activity, thus amplifying the initial mitochondrial apoptotic signaling. Conclusion All these results suggest that maslinic acid induces apoptosis in human HT29 colon-cancer cells through the JNK-Bid-mediated mitochondrial apoptotic

  15. Effect of caspase-dependent pathway on apoptosis of human gingival fibroblasts induced by advanced glycation end products%凋亡蛋白酶依赖性凋亡途径在晚期糖基化终产物诱导人牙龈成纤维细胞凋亡中的作用

    Institute of Scientific and Technical Information of China (English)

    刘开蕾; 俞少杰; 付云

    2011-01-01

    Objective To investigate the effect of synthesized advanced glycation end products (AGE) on apoptosis of human gingival fibroblasts(HGF) and the possible role of caspase-dependent pathway in the process of AGE-induced apoptosis.Methods HGF were incubated with AGE-human serum albumin (AGE-HSA).The activity of caspase-8,caspase-9 and caspase-3 were detected by microplate reader after 12 and 24 hours.HGF were incubated with caspase inhibitors for 1 hour,and then incubated with AGE-HSA for 24 hours,HGF was first stained by Hoechst33258 and observed under inverted microscope,and then double stained by annexin V and propidine iodide(PI) and observed by flow cytometry(FCM).The activity of caspase-3 was determined by caspase-3 assay kit and observed by microplate reader.Results Caspases activity of caspase-8,-9,-3 was 0.1097 ±0.0051,0.0965 ±0.0051 and 0.1280 ±0.0103 after 12 h of incubation with AGE-HSA and HGF,respectively,and 0.1558 ±0.0053,0.1308 ±0.0035 and 0.1954 ±0.0051 after 24 h of incubation with AGE-HSA and HGF,respectively ( P <0.05).Positive cells number was 247.7 ±32.4,200.1 ± 14.6,154.1 ± 14.4 and 131.3 ± 14.6 in caspase inhibitor groups by Hoechst33258 staining,respectively.Apoptotic rate was ( 25.57 ± 2.20 ) %,( 38.87 ± 3.31 ) %,( 17.17 ± 2.24 ) % and ( 14.73 ±2.48)% in caspase inhibitor groups by annexin V-PI staining,respectively.The difference between different groups was significant (P < 0.05 ).Caspase-3 activity was reduced to 0.1274 ± 0.0076,0.1465 ± 0.0062,0.1044 ±0.0051 in caspase inhibitor groups,respectively.The difference between different groups was significant(P < 0.05 ).Conclusions Apoptosis of HGF induced by AGE-HISA may be mainly through activation of caspase-dependant pathway in which cytoplasmic pathway may play a predominant role.%目的 观察晚期糖基化终产物(advanced glycation end products,AGE)对人牙龈成纤维细胞(human gingival fibroblasts,HGF)凋亡的诱导作用,同时通过检测胱天蛋白酶(caspase

  16. Full Length Bid is sufficient to induce apoptosis of cultured rat hippocampal neurons

    Directory of Open Access Journals (Sweden)

    Ward Manus W

    2007-02-01

    Full Text Available Abstract Background Bcl-2 homology domain (BH 3-only proteins are pro-apoptotic proteins of the Bcl-2 family that couple stress signals to the mitochondrial cell death pathways. The BH3-only protein Bid can be activated in response to death receptor activation via caspase 8-mediated cleavage into a truncated protein (tBid, which subsequently translocates to mitochondria and induces the release of cytochrome-C. Using a single-cell imaging approach of Bid cleavage and translocation during apoptosis, we have recently demonstrated that, in contrast to death receptor-induced apoptosis, caspase-independent excitotoxic apoptosis involves a translocation of full length Bid (FL-Bid from the cytosol to mitochondria. We induced a delayed excitotoxic cell death in cultured rat hippocampal neurons by a 5-min exposure to the glutamate receptor agonist N-methyl-D-aspartate (NMDA; 300 μM. Results Western blot experiments confirmed a translocation of FL-Bid to the mitochondria during excitotoxic apoptosis that was associated with the release of cytochrome-C from mitochondria. These results were confirmed by immunofluorescence analysis of Bid translocation during excitotoxic cell death using an antibody raised against the amino acids 1–58 of mouse Bid that is not able to detect tBid. Finally, inducible overexpression of FL-Bid or a Bid mutant that can not be cleaved by caspase-8 was sufficient to induce apoptosis in the hippocampal neuron cultures. Conclusion Our data suggest that translocation of FL-Bid is sufficient for the activation of mitochondrial cell death pathways in response to glutamate receptor overactivation.

  17. The proteins (12 and 15 kDa) isolated from heat-killed Lactobacillus plantarum L67 induces apoptosis in HT-29 cells.

    Science.gov (United States)

    Song, S; Oh, S; Lim, K T

    2015-03-01

    A number of scientific studies have revealed that Lactobacillus strains have beneficial bioactivities in the gastrointestinal tract. In this study, the production of intracellular reactive oxygen species (ROS) and the amounts of intracellular calcium, protein kinase C activity, cytochrome c, Bid, Bcl-2, Bax and the apoptosis-mediated proteins [caspase-8, caspase-3 and poly ADP ribose polymerase (PARP)] were evaluated to understand the induction of programmed cell death in HT-29 cells by Lactobacillus plantarum L67. The results obtained from this study indicated that the relative intensities of the apoptotic-related factors (intracellular ROS and intracellular calcium) and of apoptotic signals (Bax and t-Bid) increased with increasing concentrations of the membrane proteins isolated from heat-killed L. plantarum L67, whereas the relative intensities of cytochrome c, Bcl-2, caspase-8, caspase-3 and PARP decreased. This study determines whether proteins (12 and 15 kDa) isolated from heat-killed L. plantarum L67 induce programmed cell death in HT-29 cells. Proteins isolated from L. plantarum L67 can stimulate the apoptotic signals and then consequently induce programmed cell death in HT-29 cells. The results in this study suggest that the proteins isolated from L. plantarum L67 could be used as an antitumoural agent in probiotics and as a component of supplements or health foods.

  18. A novel cryptic binding motif, LRSKSRSFQVSDEQY, in the C-terminal fragment of MMP-3/7-cleaved osteopontin as a novel ligand for α9β1 integrin is involved in the anti-type II collagen antibody-induced arthritis.

    Directory of Open Access Journals (Sweden)

    Shigeyuki Kon

    Full Text Available Osteopontin (OPN is a multifunctional protein that has been linked to various intractable inflammatory diseases. One way by which OPN induces inflammation is the production of various functional fragments by enzyme cleavage. It has been well appreciated that OPN is cleaved by thrombin, and/or matrix metalloproteinase-3 and -7 (MMP-3/7. Although the function of thrombin-cleaved OPN is well characterized, little is known about the function of MMP-3/7-cleaved OPN. In this study, we found a novel motif, LRSKSRSFQVSDEQY, in the C-terminal fragment of MMP-3/7-cleaved mouse OPN binds to α9β1 integrin. Importantly, this novel motif is involved in the development of anti-type II collagen antibody-induced arthritis (CAIA. This study provides the first in vitro and in vivo evidence that OPN cleavage by MMP-3/7 is an important regulatory mechanism for CAIA.

  19. Friedreich's ataxia--a case of aberrant transcription termination?

    Science.gov (United States)

    Butler, Jill Sergesketter; Napierala, Marek

    2015-01-01

    Reduced expression of the mitochondrial protein Frataxin (FXN) is the underlying cause of Friedreich's ataxia. We propose a model of premature termination of FXN transcription induced by pathogenic expanded GAA repeats that links R-loop structures, antisense transcription, and heterochromatin formation as a novel mechanism of transcriptional repression in Friedreich's ataxia.

  20. The small molecule calactin induces DNA damage and apoptosis in human leukemia cells.

    Science.gov (United States)

    Lee, Chien-Chih; Lin, Yi-Hsiung; Chang, Wen-Hsin; Wu, Yang-Chang; Chang, Jan-Gowth

    2012-09-01

    We purified calactin from the roots of the Chinese herb Asclepias curassavica L. and analyzed its biologic effects in human leukemia cells. Our results showed that calactin treatment caused DNA damage and resulted in apoptosis. Increased phosphorylation levels of Chk2 and H2AX were observed and were reversed by the DNA damage inhibitor caffeine in calactin-treated cells. In addition, calactin treatment showed that a decrease in the expression of cell cycle regulatory proteins Cyclin B1, Cdk1, and Cdc25C was consistent with a G2/M phase arrest. Furthermore, calactin induced extracellular signal-regulated kinase (ERK) phosphorylation, activation of caspase-3, caspase-8, and caspase-9, and PARP cleavage. Pretreatment with the ERK inhibitor PD98059 significantly blocked the loss of viability in calactin-treated cells. It is indicated that calactin-induced apoptosis may occur through an ERK signaling pathway. Our data suggest that calactin is a potential anticancer compound.

  1. Action potential broadening and frequency-dependent facilitation of calcium signals in pituitary nerve terminals.

    OpenAIRE

    Jackson, M B; Konnerth, A.; Augustine, G.J.

    1991-01-01

    Hormone release from nerve terminals in the neurohypophysis is a sensitive function of action potential frequency. We have investigated the cellular mechanisms responsible for this frequency-dependent facilitation by combining patch clamp and fluorimetric Ca2+ measurements in single neurosecretory terminals in thin slices of the rat posterior pituitary. In these terminals both action potential-induced changes in the intracellular Ca2+ concentration ([Ca2+]i) and action potential duration were...

  2. Sulforaphane Attenuates Gentamicin-Induced Nephrotoxicity: Role of Mitochondrial Protection

    Science.gov (United States)

    Huerta-Yepez, Sara; Medina-Campos, Omar Noel; Zatarain-Barrón, Zyanya Lucía; Hernández-Pando, Rogelio; Torres, Ismael; Tapia, Edilia; Pedraza-Chaverri, José

    2013-01-01

    Sulforaphane (SFN), an isothiocyanate naturally occurring in Cruciferae, induces cytoprotection in several tissues. Its protective effect has been associated with its ability to induce cytoprotective enzymes through an Nrf2-dependent pathway. Gentamicin (GM) is a widely used antibiotic; nephrotoxicity is the main side effect of this compound. In this study, it was investigated if SFN is able to induce protection against GM-induced nephropathy both in renal epithelial LLC-PK1 cells in culture and in rats. SFN prevented GM-induced death and loss of mitochondrial membrane potential in LLC-PK1 cells. In addition, it attenuated GM-induced renal injury (proteinuria, increases in serum creatinine, in blood urea nitrogen, and in urinary excretion on N-acetyl-β-D-glucosaminidase, and decrease in creatinine clearance and in plasma glutathione peroxidase activity) and necrosis and apoptosis in rats. The apoptotic death was associated with enhanced active caspase-9. Caspase-8 was unchanged in all the studied groups. In addition, SFN was able to prevent GM-induced protein nitration and decrease in the activity of antioxidant enzymes catalase and glutathione peroxidase in renal cortex. In conclusion, the protective effect of SFN against GM-induced acute kidney injury could be associated with the preservation in mitochondrial function that would prevent the intrinsic apoptosis and nitrosative stress. PMID:23662110

  3. Sulforaphane Attenuates Gentamicin-Induced Nephrotoxicity: Role of Mitochondrial Protection

    Directory of Open Access Journals (Sweden)

    Mario Negrette-Guzmán

    2013-01-01

    Full Text Available Sulforaphane (SFN, an isothiocyanate naturally occurring in Cruciferae, induces cytoprotection in several tissues. Its protective effect has been associated with its ability to induce cytoprotective enzymes through an Nrf2-dependent pathway. Gentamicin (GM is a widely used antibiotic; nephrotoxicity is the main side effect of this compound. In this study, it was investigated if SFN is able to induce protection against GM-induced nephropathy both in renal epithelial LLC-PK1 cells in culture and in rats. SFN prevented GM-induced death and loss of mitochondrial membrane potential in LLC-PK1 cells. In addition, it attenuated GM-induced renal injury (proteinuria, increases in serum creatinine, in blood urea nitrogen, and in urinary excretion on N-acetyl-β-D-glucosaminidase, and decrease in creatinine clearance and in plasma glutathione peroxidase activity and necrosis and apoptosis in rats. The apoptotic death was associated with enhanced active caspase-9. Caspase-8 was unchanged in all the studied groups. In addition, SFN was able to prevent GM-induced protein nitration and decrease in the activity of antioxidant enzymes catalase and glutathione peroxidase in renal cortex. In conclusion, the protective effect of SFN against GM-induced acute kidney injury could be associated with the preservation in mitochondrial function that would prevent the intrinsic apoptosis and nitrosative stress.

  4. Conformal mapping for multiple terminals

    Science.gov (United States)

    Wang, Weimin; Ma, Wenying; Wang, Qiang; Ren, Hao

    2016-11-01

    Conformal mapping is an important mathematical tool that can be used to solve various physical and engineering problems in many fields, including electrostatics, fluid mechanics, classical mechanics, and transformation optics. It is an accurate and convenient way to solve problems involving two terminals. However, when faced with problems involving three or more terminals, which are more common in practical applications, existing conformal mapping methods apply assumptions or approximations. A general exact method does not exist for a structure with an arbitrary number of terminals. This study presents a conformal mapping method for multiple terminals. Through an accurate analysis of boundary conditions, additional terminals or boundaries are folded into the inner part of a mapped region. The method is applied to several typical situations, and the calculation process is described for two examples of an electrostatic actuator with three electrodes and of a light beam splitter with three ports. Compared with previously reported results, the solutions for the two examples based on our method are more precise and general. The proposed method is helpful in promoting the application of conformal mapping in analysis of practical problems.

  5. Ginsenoside Rh2 inhibits proliferation and induces apoptosis in human leukemia cells via TNF-α signaling pathway.

    Science.gov (United States)

    Huang, Jingjia; Peng, Kunjian; Wang, Linghao; Wen, Bin; Zhou, Lin; Luo, Tiao; Su, Min; Li, Jijia; Luo, Zhiyong

    2016-08-01

    Ginsenoside Rh2, a triterpene saponin extracted from Panax ginseng, exhibits pharmacological activity against multiple cancers. However, the anticancer mechanism of ginsenoside Rh2 is unclear. In this study, we found that ginsenoside Rh2 effectively inhibits growth and induces apoptosis of HL-60 cells. Using microarray technology, we found that tumor necrosis factor-α (TNF-α) is clearly up-regulated. Furthermore, anti-TNF-α antibody relieved the Rh2-induced HL-60 cell apoptosis via suppression of caspase-8, caspase-9, and caspase-3 activation. In addition, TNF-α up-regulation was also observed in other Rh2-treated cancer cell lines. These results demonstrate that TNF-α plays a key role in ginsenoside Rh2-induced cell apoptosis.

  6. 冬凌草甲素对骨肉瘤细胞增殖抑制和凋亡诱导效应的机制研究%Molecular Mechanism of Oridonin in Inhibiting Proliferation and Inducing Apoptosis of Osteosarcoma Cell MG-63

    Institute of Scientific and Technical Information of China (English)

    唐新桥; 朱宝玉; 王万春

    2013-01-01

    目的 研究冬凌草甲素对人骨肉瘤细胞株MG-63细胞增殖抑制和凋亡诱导效应的分子机制.方法 以流式细胞仪检测细胞周期变化;Western印迹检测CyclinD1、P21以及凋亡相关蛋白Survivin、Bcl-2和Bax表达的变化;TRAP-PCR-ELISA方法检测MG-63细胞端粒酶活性的变化;Z-IETD-fmk阻断试验检测Caspase-8阻断前后,冬凌草甲素对MG-63细胞凋亡影响的变化.结果 冬凌草甲素增加G0/G1期MG-63细胞百分率,降低S期MG-63细胞百分率;浓度依赖性抑制MG-63细胞的CyclinD1、Survivin和Bcl-2蛋白表达,上调P21和Bax蛋白表达;浓度依赖性抑制MG-63细胞端粒酶的活性;Z-IETD-fmk阻断Caspase-8活性后,能部分逆转和减弱冬凌草甲素对MG-63细胞的凋亡诱导作用.结论 冬凌草甲素通过影响细胞周期调节蛋白、阻断细胞周期G1/S期“稽查点”;抑制Survivin、Bcl-2,上调Bax;抑制端粒酶活性以及活化Caspase-8等途径抑制MG-63细胞增殖及诱导MG-63细胞凋亡.%Objective To study the molecular mechanism of oridonin in the inhibiting the proliferation and inducing apoptosis of osteosarcoma cell MG-63. Methods The change of cell cycle was analyzed by flow cytometry, the protein expression of CyclinD1, P21, and apoptosis-associated protein Survivin, Bcl-2 and Bax in MG-63 cells were detected by Western blotting method. TRAP-PCR-ELISA was applied for the detection of telomerase activities. After the activation of Caspase-8 was inhibited, the proliferation-inhibiting and apoptosis-inducing effects of Oridonin were detected by Z-IETD-fmk blocking test. Results Oridonin increased the percentage of G0/G1 phase and decreased S phase of MG-63 cells, down-regulated CyclinD1, Survivin and Bcl-2, and up-regulated P21 and Bax protein in a concentration-dependent way in MG-63 cells. Oridonin down-regulated telomerase activities in a concentration-dependent way in MG-63 cells. The apoptosis-inducing effects of oridonin were partly

  7. Digital autonomous terminal access communications

    Science.gov (United States)

    Novacki, S.

    1987-01-01

    A significant problem for the Bus Monitor Unit is to identify the source of a given transmission. This problem arises from the fact that the label which identifies the source of the transmission as it is put into the bus is intercepted by the Digital Autonomous Terminal Access Communications (DATAC) terminal and removed from the transmission. Thus, a given subsystem will see only data associated with a label and never the identifying label itself. The Bus Monitor must identify the source of the transmission so as to be able to provide some type of error identification/location in the event that some problem with the data transmission occurs. Steps taken to alleviate this problem by modifications to the DATAC terminal are discussed.

  8. Termination of Nondeterministic Quantum Programs

    CERN Document Server

    Li, Yangjia; Ying, Mingsheng

    2012-01-01

    We define a language-independent model of nondeterministic quantum programs in which a quantum program consists of a finite set of quantum processes. These processes are represented by quantum Markov chains over the common state space. An execution of a nondeterministic quantum program is modeled by a sequence of actions of individual processes. These actions are described by super-operators on the state Hilbert space. At each step of an execution, a process is chosen nondeterministically to perform the next action. A characterization of reachable space and a characterization of diverging states of a nondeterministic quantum program are presented. We establish a zero-one law for termination probability of the states in the reachable space of a nondeterministic quantum program. A combination of these results leads to a necessary and sufficient condition for termination of nondeterministic quantum programs. Based on this condition, an algorithm is found for checking termination of nondeterministic quantum progr...

  9. 12 CFR 611.1220 - Termination resolution.

    Science.gov (United States)

    2010-01-01

    ... 12 Banks and Banking 6 2010-01-01 2010-01-01 false Termination resolution. 611.1220 Section 611... Institution Status § 611.1220 Termination resolution. No more than 1 week before you submit your plan of termination to us, your board of directors must adopt a termination resolution stating its support...

  10. Operational Semantics of Termination Types

    DEFF Research Database (Denmark)

    Nielson, Flemming; Nielson, Hanne Riis

    1996-01-01

    In principle termination analysis is easy: find a well-founded ordering and prove that calls decrease with respect to the ordering. We show how to embed termination information into a polymorphic type system for an eager higher-order functional language allowing multiple-argument functions...... and algebraic data types. The well-founded orderings are defined by pattern matching against the definition of the algebraic data types. We prove that the analysis is semantically sound with respect to a big-step (or natural) operational semantics. We compare our approach based on operational semantics to one...

  11. Stream Productivity by Outermost Termination

    CERN Document Server

    Zantema, Hans; 10.4204/EPTCS.15.7

    2010-01-01

    Streams are infinite sequences over a given data type. A stream specification is a set of equations intended to define a stream. A core property is productivity: unfolding the equations produces the intended stream in the limit. In this paper we show that productivity is equivalent to termination with respect to the balanced outermost strategy of a TRS obtained by adding an additional rule. For specifications not involving branching symbols balancedness is obtained for free, by which tools for proving outermost termination can be used to prove productivity fully automatically.

  12. Combining norms to prove termination

    DEFF Research Database (Denmark)

    Genaim, S.; Codish, M.; Gallagher, John Patrick;

    2002-01-01

    of deriving automatically a candidate norm with which to prove termination. Instead of deriving a single, complex norm function, it is sufficient to determine a collection of simpler norms, some combination of which, leads to a proof of termination. We propose that a collection of simple norms, one for each...... of the recursive data-types in the program, is often a suitable choice. We first demonstrate the power of combining norm functions and then the adequacy of combining norms based on regular types....

  13. Application of RU 486 Combined with Prostaglandin Analogues forTermination of Early Pregnancy in China

    Institute of Scientific and Technical Information of China (English)

    俊康; 朱月华; 贺昌海; 宋思; 何美丽; 杨秋英

    1992-01-01

    The induced abortion carried out in China is mainly due to contraceptive faiture and considered to be an important comptementay, measure . In China the vacuum aspiration have been used.for termination of early pregnancy since 1958. Althrough thecomplication rate of this procedure is very low, surgical termination of pregnancy re-quires experienced health care personnels.

  14. Enrichment and terminal differentiation of striated muscle progenitors in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Becher, Ulrich M.; Breitbach, Martin; Sasse, Philipp [Institute of Physiology I, Life and Brain Center, University of Bonn, Bonn (Germany); Garbe, Stephan [Department of Radiology, University of Bonn, Bonn (Germany); Ven, Peter F.M. van der [Institute for Cell Biology, University of Bonn, Bonn (Germany); Fuerst, Dieter O., E-mail: dfuerst@uni-bonn.de [Institute for Cell Biology, University of Bonn, Bonn (Germany); Fleischmann, Bernd K., E-mail: bernd.fleischmann@uni-bonn.de [Institute of Physiology I, Life and Brain Center, University of Bonn, Bonn (Germany)

    2009-10-01

    Enrichment and terminal differentiation of mammalian striated muscle cells is severely hampered by fibroblast overgrowth, de-differentiation and/or lack of functional differentiation. Herein we report a new, reproducible and simple method to enrich and terminally differentiate muscle stem cells and progenitors from mice and humans. We show that a single gamma irradiation of muscle cells induces their massive differentiation into structurally and functionally intact myotubes and cardiomyocytes and that these cells can be kept in culture for many weeks. Similar results are also obtained when treating skeletal muscle-derived stem cells and progenitors with Mitomycin C.

  15. Spontaneous and field-induced mesomorphism of a silyl-terminated bent-core liquid crystal as determined from second-harmonic generation and resonant X-ray scattering.

    Science.gov (United States)

    Folcia, C L; Ortega, J; Etxebarria, J; Rodríguez-Conde, S; Sanz-Enguita, G; Geese, K; Tschierske, C; Ponsinet, V; Barois, P; Pindak, R; Pan, LiDong; Liu, Z Q; McCoy, B K; Huang, C C

    2014-01-07

    The polarity and structure of the phases of a liquid crystal constituted by thiophene-based bent-core molecules is investigated by means of optical second-harmonic generation (SHG), and resonant and conventional X-ray diffraction. The material studied is representative of a wide family of mesogens that contain silyl groups at the ends of the chains. These bulky terminal groups have been reported to give rise to smectic phases showing ferroelectric switching. However, the analysis of the SHG signal before and after application of electric fields has allowed us to establish unambiguously that the reported ferroelectricity is not intrinsic to the material but stabilized by the cell substrates once an electric field has been applied. In addition, the results obtained from resonant X-ray diffraction indicate that virgin samples have antiferroelectric undulated synclinic smectic structures.

  16. The impact of 'terminator' technology

    NARCIS (Netherlands)

    Visser, B.; Meer, van der I.J.M.; Louwaars, N.; Beekwilder, J.; Eaton, D.

    2001-01-01

    Genetic use-restriction technologies enable the developers of transgenic plants or animals to protect their variety or breed from unauthorized use in a biological way. The use of 'terminator technology' can have different impacts on farmers and breeders. If the technology is effective, it impacts on

  17. What Determines Joint Venture Termination?

    DEFF Research Database (Denmark)

    Nielsen, Bo Bernhard

    2012-01-01

    Joint venture (JV) research continues to flourish as researchers seek to advance our understanding of why so many JVs fail. Cui and Kumar (this issue) take a contingency approach to explain how and why business relatedness may provide new insights as to what determines JV termination...

  18. Equiseparability on Terminal Wiener Index

    DEFF Research Database (Denmark)

    Deng, Xiaotie; Zhang, Jie

    2012-01-01

    The aim of this work is to explore the properties of the terminal Wiener index, which was recently proposed by Gutman et al. (2004) [3], and to show the fact that there exist pairs of trees and chemical trees which cannot be distinguished by using it. We give some general methods for constructing...

  19. 芍药苷抑制FasL诱导的Fas/FasL信号转导通路的实验研究%Experimental Research of Paeoniflorin Inhibition on Fas/FasL Signal Transduction Pathway Induced by FasL

    Institute of Scientific and Technical Information of China (English)

    陈少清; 林建平; 王诗忠; 廖军

    2012-01-01

    Objective To explore the impacts of paeoniflorin on Fas/Fasl signal transduction pathway of the cervical disc fiber cells induced by Fasl in the rats. Methods The cervical intervertebral disc an-nulus fibrosus was collected from SD rats( either sex )of 1 month old. The enzyme digestion was adopted to establish the in vitro culture of intervertebral disc annulus fibrosus cells. When the annulus fibrosus cells were passed down till the 3rd generation, the apoptosis model of the in vitro annulus fibrosus cells induced by FasL was set up. In the experiment, the cells were randomized into a blank group, a model group and a medicine group. In the model group, FasL of the proper concentration was used to induce apoptosis. In the medicine group, at the same time of Fasl induction, paeoniflorin culture solution of a proper concentration was given for the intervention. Annexin V/FITC was adopted to detect the apoptosis rate, identify the model duplication and observe the protection of paeoniflorin. The fluorescence quantitative polymerase chain reaction( FQ - PCR ) technique was used to detect the expression levels of Fas, Caspase - 3 and Caspase - 8. Results In the medicine group, the apoptosis rate of annulus fibrosus cells was lower apparently than that in the model group( P < 0. 01 ). In the medicine group, the expressions of apoptotic factor genes as Fas, Caspase - 3 and Caspase -8 were lower obviously than those in the model group( P < 0.01 ). Conclusion The paeoniflorin inhibits the transduction pathway of Fas/FasL signals through the down - regulation of the genetic expressions of Fas, Caspase - 3 and Caspase - 8 of annulus fibrosus cells so that the apoptosis rate of annulus fibrosus cells can be decreased.%目的 探讨芍药苷对Fas配体(FasL)诱导大鼠颈椎间盘纤维细胞的Fas/FasL信号转导通路的影响.方法 采用1月龄SD大鼠(雌雄不拘)的颈椎间盘纤维环,采用酶消化法,建立椎间盘纤维环细胞体外培养.传至3代纤维

  20. 纳秒脉冲诱导SKOV3细胞凋亡的死亡受体途径分析%Analysis on Death Receptor Apoptotic Pathway of SKOV3 Cells Induced by Nanosecond Pulsed Electric Field

    Institute of Scientific and Technical Information of China (English)

    郭飞; 姚陈果; 王建; 孙才新; 夏如民; 唐均英

    2012-01-01

    The specific bioelectric effect of tumor cells apoptosis induced by nanosecond pulsed electric field ( nsPEF) has aroused great attention. Based on the latest studies, the effects of nsPEF on plasma membrane were illustrated to study the signaling pathway of death receptor apoptotic. Therefore, optimized parameters (voltage amplitude of 9 kV, pulse duration of 100 ns, pulse number of 30, repetition frequency of 1 Hz) of nsPEF were performed on SKOVa cells. Cell death and apoptosis were tested by flow cytometry, massager ribonucleic acid[mRNA) release of Fas, FasL, cysteine aspartic acid specific protease-8 (Caspase 8) and Bid were examined by reverse transcription-polymerase chain reaction(RT-PCR) method, and protein release of Fas, FasL, Caspase-8 and Bid were studied by western blot technology. Experimental results indicate that release of Fas, FasL, Caspase-8 and Bid greatly increase when tumor cell apoptosis with nsPEF, in advance to trigger the apoptotic signaling pathway of death receptor. The results provide a theoretic support for clinical tumor treatment with boarding the mechanism study of nsPEF-indueed apoptosis.%ns脉冲电场独特的诱导肿瘤细胞凋亡的生物电效应,引起了相关学者广泛的关注。为此,结合最新研究成果,侧重于ns脉冲对细胞膜结构和功能的影响,重点研究ns脉冲电场诱导肿瘤细胞凋亡的死亡受体途径。将优化的脉冲电场参数组合(电压幅值为9kV,脉宽为100ns,脉冲为30个,频率为1Hz)作用于人卵巢浆液性囊腺癌细胞SKOV3。利用流式细胞术和凝胶电泳法检测细胞凋亡、坏死情况;逆转录聚合酶链式扩增反应(reverse transcription-polymerase chain reaction,RT-PCR)法检测Fas、FasL、半胱氨酸天冬氨酸蛋白酶-8(cysteine aspartic acid specific protease-8,Caspase-8)和Bid的信使核糖核酸(messager ribonucleic acid,mRNA)释放水平;蛋白质印迹(western blot)法检测Fas、FasL、Caspase

  1. Twist haploinsufficiency in Saethre-Chotzen syndrome induces calvarial osteoblast apoptosis due to increased TNFalpha expression and caspase-2 activation.

    Science.gov (United States)

    Yousfi, Malika; Lasmoles, Francoise; El Ghouzzi, Vincent; Marie, Pierre J

    2002-02-15

    Saethre-Chotzen syndrome (SCS) is a human autosomal dominant disorder characterized by premature fusion of cranial sutures caused by mutations of the Twist gene encoding a basic helix-loop-helix (bHLH) transcription factor. We previously showed that Twist haploinsufficiency caused by a Y103X nonsense mutation in SCS alters both proliferation and osteoblast gene expression in human calvarial osteoblasts, indicating that Twist is an important regulator of osteoblast differentiation. Here we show that Twist haploinsufficiency alters osteoblast apoptosis in SCS. Analysis of terminal deoxynucleotidyl transferase-mediated nick-end labelling (TUNEL) demonstrated increased osteoblast and osteocyte apoptosis in coronal sutures from two SCS patients with nonsense mutations (Y103X and Q109X) that result in the synthesis of bHLH-truncated proteins, and one patient with a missense mutation in the basic domain (R118C) that abolishes Twist DNA binding. To assess the mechanisms involved, we studied osteoblast apoptosis in mutant (M-Tw) calvarial cells bearing the Y103X mutation resulting in decreased Twist mRNA and protein levels. M-Tw cells cultured in low serum conditions showed enhanced DNA fragmentation compared to normal (Nl) age-matched calvarial cells. Biochemical analysis showed increased activity of initiator caspases-2 and -8 and downstream effector caspases-3, -6 and -7 in mutant osteoblasts. Caspase-2 was upstream of caspase-8 and effector caspases-3, -6 and -7 because their activities were suppressed by a specific caspase-2 inhibitor. M-Tw osteoblasts also showed increased cytochrome c release from the mitochondria. However, the activity of the downstream effector caspase-9 was not increased due to overexpression of the antagonist protein Hsp70. Detection of differentially expressed genes using cDNA expression array revealed increased Bax and TNFalpha mRNA levels in M-Tw compared to Nl cells, a finding confirmed by RT-PCR and western blot analyses. Neutralization of

  2. Development of Noviomimetics as C-Terminal Hsp90 Inhibitors.

    Science.gov (United States)

    Anyika, Mercy; McMullen, Mason; Forsberg, Leah K; Dobrowsky, Rick T; Blagg, Brian S J

    2016-01-14

    KU-32 and KU-596 are novobiocin-derived, C-terminal heat shock protein 90 (Hsp90) modulators that induce Hsp70 levels and manifest neuroprotective activity. However, the synthetically complex noviose sugar requires 10 steps to prepare, which makes translational development difficult. In this study, we developed a series of "noviomimetic" analogues of KU-596, which contain noviose surrogates that can be easily prepared, while maintaining the ability to induce Hsp70 levels. Both sugar and sugar analogues were designed, synthesized, and evaluated in a luciferase reporter assay, which identified compound 37, a benzyl containing noviomimetic, as the most potent inducer of Hsp70.

  3. Singlet oxygen treatment of tumor cells triggers extracellular singlet oxygen generation, catalase inactivation and reactivation of intercellular apoptosis-inducing signaling.

    Science.gov (United States)

    Riethmüller, Michaela; Burger, Nils; Bauer, Georg

    2015-12-01

    Intracellular singlet oxygen generation in photofrin-loaded cells caused cell death without discrimination between nonmalignant and malignant cells. In contrast, extracellular singlet oxygen generation caused apoptosis induction selectively in tumor cells through singlet oxygen-mediated inactivation of tumor cell protective catalase and subsequent reactivation of intercellular ROS-mediated apoptosis signaling through the HOCl and the NO/peroxynitrite signaling pathway. Singlet oxygen generation by extracellular photofrin alone was, however, not sufficient for optimal direct inactivation of catalase, but needed to trigger the generation of cell-derived extracellular singlet oxygen through the interaction between H2O2 and peroxynitrite. Thereby, formation of peroxynitrous acid, generation of hydroxyl radicals and formation of perhydroxyl radicals (HO2(.)) through hydroxyl radical/H2O2 interaction seemed to be required as intermediate steps. This amplificatory mechanism led to the formation of singlet oxygen at a sufficiently high concentration for optimal inactivation of membrane-associated catalase. At low initial concentrations of singlet oxygen, an additional amplification step needed to be activated. It depended on singlet oxygen-dependent activation of the FAS receptor and caspase-8, followed by caspase-8-mediated enhancement of NOX activity. The biochemical mechanisms described here might be considered as promising principle for the development of novel approaches in tumor therapy that specifically direct membrane-associated catalase of tumor cells and thus utilize tumor cell-specific apoptosis-inducing ROS signaling.

  4. Termination of pregnancy under French law: from criminalization to a right in accordance with international developments on women's rights.

    Science.gov (United States)

    Madanamoothoo, Allane

    2011-12-01

    Termination of pregnancy is the premature exit of the products of conception, which include the placenta, bag of waters, embryo or fetus from the uterus. In general, the term "termination of pregnancy" refers to non-medical termination of pregnancy, which is requested for different reasons other than medical ones. When such a request is made in countries where it is lawful, women have access to induced termination of pregnancy under lawful and limited conditions. However, in countries where the practice is illegal, women tend to suffer and die of complications from unsafe termination of pregnancy. Nowadays, there seems to be a worldwide trend towards the legalization of termination of pregnancy. The impact of international developments on women's rights has played an increasing role in improving access to termination of pregnancy. This article aims at describing how legalization of termination of pregnancy in France has become a right which is in accordance with international developments on women's rights.

  5. Mobile Terminal Management in 3G

    Institute of Scientific and Technical Information of China (English)

    SHANG Jing; MAN Yi; SONG Jun-de; SONG Mei-na

    2005-01-01

    Recently, Terminal has become one of the key issues in the 3G development. In the paper, we study the mobile terminal management in 3G network. Based on the analysis of problems encountered with the terminals, the paper describes the use cases of terminal management. A system architecture with its function components of mobile terminal management system is proposed. Moreover, a possible implementation solution is introduced and described. A Management Information Model (MIM) for terminal management is also proposed to support the management services.

  6. 1-Benzyl-2-Phenylbenzimidazole (BPB, a Benzimidazole Derivative, Induces Cell Apoptosis in Human Chondrosarcoma through Intrinsic and Extrinsic Pathways

    Directory of Open Access Journals (Sweden)

    Ju-Fang Liu

    2012-12-01

    Full Text Available In this study, we investigated the anticancer effects of a new benzimidazole derivative, 1-benzyl-2-phenyl -benzimidazole (BPB, in human chondrosarcoma cells. BPB-mediated apoptosis was assessed by the MTT assay and flow cytometry analysis. The in vivo efficacy was examined in a JJ012 xenograft model. Here we found that BPB induced apoptosis in human chondrosarcoma cell lines (JJ012 and SW1353 but not in primary chondrocytes. BPB induced upregulation of Bax, Bad and Bak, downregulation of Bcl-2, Bid and Bcl-XL and dysfunction of mitochondria in chondrosarcoma. In addition, BPB also promoted cytosolic releases AIF and Endo G. Furthermore, it triggered extrinsic death receptor-dependent pathway, which was characterized by activating Fas, FADD and caspase-8. Most importantly, animal studies revealed a dramatic 40% reduction in tumor volume after 21 days of treatment. Thus, BPB may be a novel anticancer agent for the treatment of chondrosarcoma.

  7. Tumor necrosis factor (TNF) receptor-associated factor 7 is required for TNFα-induced Jun NH2-terminal kinase activation and promotes cell death by regulating polyubiquitination and lysosomal degradation of c-FLIP protein.

    Science.gov (United States)

    Scudiero, Ivan; Zotti, Tiziana; Ferravante, Angela; Vessichelli, Mariangela; Reale, Carla; Masone, Maria C; Leonardi, Antonio; Vito, Pasquale; Stilo, Romania

    2012-02-17

    The pro-inflammatory cytokine tumor necrosis factor (TNF) α signals both cell survival and death. The biological outcome of TNFα treatment is determined by the balance between survival factors and Jun NH(2)-terminal kinase (JNK) signaling, which promotes cell death. Here, we show that TRAF7, the most recently identified member of the TNF receptor-associated factors (TRAFs) family of proteins, is essential for activation of JNK following TNFα stimulation. We also show that TRAF6 and TRAF7 promote unconventional polyubiquitination of the anti-apoptotic protein c-FLIP(L) and demonstrate that degradation of c-FLIP(L) also occurs through a lysosomal pathway. RNA interference-mediated depletion of TRAF7 correlates with increased c-FLIP(L) expression level, which, in turn, results in resistance to TNFα cytotoxicity. Collectively, our results indicate an important role for TRAF7 in the activation of JNK following TNFα stimulation and clearly point to an involvement of this protein in regulating the turnover of c-FLIP and, consequently, cell death.

  8. Tumor Necrosis Factor (TNF) Receptor-associated Factor 7 Is Required for TNFα-induced Jun NH2-terminal Kinase Activation and Promotes Cell Death by Regulating Polyubiquitination and Lysosomal Degradation of c-FLIP Protein*

    Science.gov (United States)

    Scudiero, Ivan; Zotti, Tiziana; Ferravante, Angela; Vessichelli, Mariangela; Reale, Carla; Masone, Maria C.; Leonardi, Antonio; Vito, Pasquale; Stilo, Romania

    2012-01-01

    The pro-inflammatory cytokine tumor necrosis factor (TNF) α signals both cell survival and death. The biological outcome of TNFα treatment is determined by the balance between survival factors and Jun NH2-terminal kinase (JNK) signaling, which promotes cell death. Here, we show that TRAF7, the most recently identified member of the TNF receptor-associated factors (TRAFs) family of proteins, is essential for activation of JNK following TNFα stimulation. We also show that TRAF6 and TRAF7 promote unconventional polyubiquitination of the anti-apoptotic protein c-FLIPL and demonstrate that degradation of c-FLIPL also occurs through a lysosomal pathway. RNA interference-mediated depletion of TRAF7 correlates with increased c-FLIPL expression level, which, in turn, results in resistance to TNFα cytotoxicity. Collectively, our results indicate an important role for TRAF7 in the activation of JNK following TNFα stimulation and clearly point to an involvement of this protein in regulating the turnover of c-FLIP and, consequently, cell death. PMID:22219201

  9. Clothes Dryer Automatic Termination Evaluation

    Energy Technology Data Exchange (ETDEWEB)

    TeGrotenhuis, Ward E.

    2014-10-01

    Volume 2: Improved Sensor and Control Designs Many residential clothes dryers on the market today provide automatic cycles that are intended to stop when the clothes are dry, as determined by the final remaining moisture content (RMC). However, testing of automatic termination cycles has shown that many dryers are susceptible to over-drying of loads, leading to excess energy consumption. In particular, tests performed using the DOE Test Procedure in Appendix D2 of 10 CFR 430 subpart B have shown that as much as 62% of the energy used in a cycle may be from over-drying. Volume 1 of this report shows an average of 20% excess energy from over-drying when running automatic cycles with various load compositions and dryer settings. Consequently, improving automatic termination sensors and algorithms has the potential for substantial energy savings in the U.S.

  10. Pentamidine sensitizes chronic myelogenous leukemia K562 cells to TRAIL-induced apoptosis.

    Science.gov (United States)

    Qiu, Geng; Jiang, Jikai; Liu, Xiao-shan

    2012-11-01

    Pentamidine (PMD) is an anti-protozoa drug with potential anticancer activity. Here we show that PMD at clinically achievable plasma drug concentrations slightly inhibited the growth of human leukemia cell lines. PMD close to its therapeutic doses sensitized TRAIL-resistant K562 cells to the cytokine and potentiated TRAIL-induced apoptosis through activation of caspase-8 and -3. When we investigated the underlying mechanism, we observed that treatment with PMD increased DR5 expression at both mRNA and protein levels and down-regulated anti-apoptotic XIAP and Mcl-1 protein levels. This study provides a rationale for a more in-depth exploration into the combined treatment with PMD and TRAIL as a valuable strategy for leukemia therapy.

  11. Phloroglucinol induces apoptosis via apoptotic signaling pathways in HT-29 colon cancer cells

    Science.gov (United States)

    KANG, MI-HYE; KIM, IN-HYE; NAM, TAEK-JEO NG

    2014-01-01

    Phloroglucinol is a polyphenolic compound that is used to treat and prevent several human diseases, as it exerts beneficial biological activities, including anti-oxidant, anti-inflammatory and anticancer properties. The aim of the present study was to investigate the effects of phloroglucinol on apoptotic signaling pathways in HT-29 colon cancer cells. The results indicated that phloroglucinol suppressed cell viability and induced apoptosis in HT-29 cells in a concentration-dependent manner. Phloroglucinol treatment of HT-29 cells resulted in characteristic apoptosis-related changes: altered Bcl-2 family proteins, cytochrome c release, and activation of caspase-3 and caspase-8. This study also showed that proteins involved in apoptosis were stimulated by treatment with phloroglucinol. These findings demonstrated that phloroglucinol exerts anticancer activity in HT-29 colon cancer cells through induction of apoptosis. PMID:25070748

  12. Investigation of the apoptotic way induced by digallic acid in human lymphoblastoid TK6 cells

    Directory of Open Access Journals (Sweden)

    Bhouri Wissem

    2012-06-01

    Full Text Available Abstract Background The digallic acid (DGA purified from Pistacia lentiscus. L fruits was investigated for its antiproliferative and apoptotic activities on human lymphoblastoid TK6 cells. Methods We attempt to characterize the apoptotic pathway activated by DGA. Apoptosis was detected by DNA fragmentation, PARP cleavage and by evaluating caspase activities. Results The inhibition of lymphoblastoid cell proliferation was noted from 8.5 μg/ml of DGA. The induction of apoptosis was confirmed by DNA fragmentation and PARP cleavage. We have demonstrated that DGA induces apoptosis by activating the caspase-8 extrinsic pathway. Caspase-3 was also activated in a dose dependent manner. Conclusion In summary, DGA exhibited an apoptosis inductor effect in TK6 cells revealing thus its potential as a cancer-preventive agent.

  13. High Octane Fuel: Terminal Backgrounder

    Energy Technology Data Exchange (ETDEWEB)

    Moriarty, Kristi [National Renewable Energy Lab. (NREL), Golden, CO (United States)

    2016-02-11

    The Bioenergy Technologies Office of the U.S. Department of Energy Office of Energy Efficiency and Renewable Energy sponsored a scoping study to assess the potential of ethanol-based high octane fuel (HOF) to reduce energy consumption and greenhouse gas emissions. When the HOF blend is made with 25%-40% ethanol by volume, this energy efficiency improvement is potentially sufficient to offset the reduced vehicle range often associated with the decreased volumetric energy density of ethanol. The purpose of this study is to assess the ability of the fuel supply chain to accommodate more ethanol at fuel terminals. Fuel terminals are midstream in the transportation fuel supply chain and serve to store and distribute fuels to end users. While there are no technical issues to storing more ethanol at fuel terminals, there are several factors that could impact the ability to deploy more ethanol. The most significant of these issues include the availability of land to add more infrastructure and accommodate more truck traffic for ethanol deliveries as well as a lengthy permitting process to erect more tanks.

  14. 48 CFR 12.403 - Termination.

    Science.gov (United States)

    2010-10-01

    ... entitled “Termination for the Government's Convenience” and “Termination for Cause” contain concepts which... required to comply with the cost accounting standards or the contract cost principles in part 31....

  15. Identification of RIP1 as a critical mediator of Smac mimetic-mediated sensitization of glioblastoma cells for Drozitumab-induced apoptosis

    Science.gov (United States)

    Cristofanon, S; Abhari, B A; Krueger, M; Tchoghandjian, A; Momma, S; Calaminus, C; Vucic, D; Pichler, B J; Fulda, S

    2015-01-01

    This study aims at evaluating the combination of the tumor-necrosis-factor-related apoptosis-inducing ligand (TRAIL)-receptor 2 (TRAIL-R2)-specific antibody Drozitumab and the Smac mimetic BV6 in preclinical glioblastoma models. To this end, the effect of BV6 and/or Drozitumab on apoptosis induction and signaling pathways was analyzed in glioblastoma cell lines, primary glioblastoma cultures and glioblastoma stem-like cells. Here, we report that BV6 and Drozitumab synergistically induce apoptosis and reduce colony formation in several glioblastoma cell lines (combination index<0.1). Also, BV6 profoundly enhances Drozitumab-induced apoptosis in primary glioblastoma cultures and glioblastoma stem-like cells. Importantly, BV6 cooperates with Drozitumab to suppress tumor growth in two glioblastoma in vivo models including an orthotopic, intracranial mouse model, underlining the clinical relevance of these findings. Mechanistic studies reveal that BV6 and Drozitumab act in concert to trigger the formation of a cytosolic receptor-interacting protein (RIP) 1/Fas-associated via death domain (FADD)/caspase-8-containing complex and subsequent activation of caspase-8 and -3. BV6- and Drozitumab-induced apoptosis is blocked by the caspase inhibitor zVAD.fmk, pointing to caspase-dependent apoptosis. RNA interference-mediated silencing of RIP1 almost completely abolishes the BV6-conferred sensitization to Drozitumab-induced apoptosis, indicating that the synergism critically depends on RIP1 expression. In contrast, both necrostatin-1, a RIP1 kinase inhibitor, and Enbrel, a TNFα-blocking antibody, do not interfere with BV6/Drozitumab-induced apoptosis, demonstrating that apoptosis occurs independently of RIP1 kinase activity or an autocrine TNFα loop. In conclusion, the rational combination of BV6 and Drozitumab presents a promising approach to trigger apoptosis in glioblastoma, which warrants further investigation. PMID:25880091

  16. 49 CFR 374.309 - Terminal facilities.

    Science.gov (United States)

    2010-10-01

    ... 49 Transportation 5 2010-10-01 2010-10-01 false Terminal facilities. 374.309 Section 374.309... REGULATIONS Adequacy of Intercity Motor Common Carrier Passenger Service § 374.309 Terminal facilities. (a... attendants and be regularly patrolled. (b) Outside facilities. At terminals and stations that are closed...

  17. 75 FR 72771 - Gap in Termination Provisions

    Science.gov (United States)

    2010-11-26

    ... specified by statute) the right to terminate certain grants of transfers or licenses within the time frames... the legislative history of the 1976 Copyright Act and the express exceptions that are found within the... termination in a timely manner is a fatal error that will prevent termination from taking effect. The...

  18. 29 CFR 405.4 - Terminal report.

    Science.gov (United States)

    2010-07-01

    ... of this part, who during its fiscal year loses its identity as a reporting employer through merger, consolidation, dissolution, or otherwise, shall, within 30 days of the effective date thereof, file a terminal...'s fiscal year ending on the effective date of the employer's termination or loss of...

  19. LNG TERMINAL SAFE OPERATION MANAGEMENT

    OpenAIRE

    Andrzej ADAMKIEWICZ; Kamiński, Włodzimierz

    2012-01-01

    W artykule przedstawiono znaczenie problemu bezpieczeństwa terminali LNG w transporcie morskim gazu ziemnego. Wskazano szczególne wymagania jakie stwarza instalacji przesył LNG, a wynikające z jego szczególnych własności. Na tle wielopoziomowych krytycznych obszarów bezpieczeństwa stanowiących elementy składowe struktury systemu bezpieczeństwa terminali wyselekcjonowano możliwości zmniejszania ryzyka wystąpienia sytuacji awaryjnej na terminalu LNG. Zdefiniowano zadania wykonywane przez termin...

  20. Resetting the transcription factor network reverses terminal chronic hepatic failure.

    Science.gov (United States)

    Nishikawa, Taichiro; Bell, Aaron; Brooks, Jenna M; Setoyama, Kentaro; Melis, Marta; Han, Bing; Fukumitsu, Ken; Handa, Kan; Tian, Jianmin; Kaestner, Klaus H; Vodovotz, Yoram; Locker, Joseph; Soto-Gutierrez, Alejandro; Fox, Ira J

    2015-04-01

    The cause of organ failure is enigmatic for many degenerative diseases, including end-stage liver disease. Here, using a CCl4-induced rat model of irreversible and fatal hepatic failure, which also exhibits terminal changes in the extracellular matrix, we demonstrated that chronic injury stably reprograms the critical balance of transcription factors and that diseased and dedifferentiated cells can be returned to normal function by re-expression of critical transcription factors, a process similar to the type of reprogramming that induces somatic cells to become pluripotent or to change their cell lineage. Forced re-expression of the transcription factor HNF4α induced expression of the other hepatocyte-expressed transcription factors; restored functionality in terminally diseased hepatocytes isolated from CCl4-treated rats; and rapidly reversed fatal liver failure in CCl4-treated animals by restoring diseased hepatocytes rather than replacing them with new hepatocytes or stem cells. Together, the results of our study indicate that disruption of the transcription factor network and cellular dedifferentiation likely mediate terminal liver failure and suggest reinstatement of this network has therapeutic potential for correcting organ failure without cell replacement.

  1. Methyl Sartortuoate Inhibits Colon Cancer Cell Growth by Inducing Apoptosis and G2/M-Phase Arrest.

    Science.gov (United States)

    Lan, Qiusheng; Li, Shoufeng; Lai, Wei; Xu, Heyang; Zhang, Yang; Zeng, Yujie; Lan, Wenjian; Chu, Zhonghua

    2015-08-17

    The potential anti-neoplastic activity of terpenoids is of continued interest. In this study, we investigate whether methyl sartortuoate, a terpenoid isolated from soft coral, induced cell cycle arrest and apoptosis in a human colon cancer cell line. Culture studies found that methyl sartortuoate inhibited colon cancer cell (LoVo and RKO) growth and caused apoptotic death in a concentration- and time-dependent manner, by activation of caspase-8, caspase-9, caspase-3, p53 and Bax, and inactivation of B-cell lymphoma 2 (Bcl-2) apoptosis regulating proteins. Methyl sartortuoate treatment led to reduced expression of cdc2 and up-regulated p21 and p53, suggesting that Methyl sartortuoate induced G2-M arrest through modulation of p53/p21/cdc2 pathways. Methyl sartortuoate also up-regulated phospho-JNK and phospho-p38 expression levels. This resulted in cell cycle arrest at the G2-M phase and apoptosis in LoVo and RKO cells. Treatment with the JNK inhibitor SP600125 and the p38 MAPK inhibitor SB203580 prevented methyl sartortuoate-induced apoptosis in LoVo cells. Moreover, methyl sartortuoate also prevented neoplasm growth in NOD-SCID nude mice inoculated with LoVo cells. Taken together, these findings suggest that methyl sartortuoate is capable of leading to activation of caspase-8, -9, -3, increasing p53 and Bax/Bcl-2 ratio apoptosis through MAPK-dependent apoptosis and results in G2-M phase arrest in LoVo and RKO cells. Thus, methyl sartortuoate may be a promising anticancer candidate.

  2. Methyl Sartortuoate Inhibits Colon Cancer Cell Growth by Inducing Apoptosis and G2/M-Phase Arrest

    Directory of Open Access Journals (Sweden)

    Qiusheng Lan

    2015-08-01

    Full Text Available The potential anti-neoplastic activity of terpenoids is of continued interest. In this study, we investigate whether methyl sartortuoate, a terpenoid isolated from soft coral, induced cell cycle arrest and apoptosis in a human colon cancer cell line. Culture studies found that methyl sartortuoate inhibited colon cancer cell (LoVo and RKO growth and caused apoptotic death in a concentration- and time-dependent manner, by activation of caspase-8, caspase-9, caspase-3, p53 and Bax, and inactivation of B-cell lymphoma 2 (Bcl-2 apoptosis regulating proteins. Methyl sartortuoate treatment led to reduced expression of cdc2 and up-regulated p21 and p53, suggesting that Methyl sartortuoate induced G2-M arrest through modulation of p53/p21/cdc2 pathways. Methyl sartortuoate also up-regulated phospho-JNK and phospho-p38 expression levels. This resulted in cell cycle arrest at the G2-M phase and apoptosis in LoVo and RKO cells. Treatment with the JNK inhibitor SP600125 and the p38 MAPK inhibitor SB203580 prevented methyl sartortuoate-induced apoptosis in LoVo cells. Moreover, methyl sartortuoate also prevented neoplasm growth in NOD-SCID nude mice inoculated with LoVo cells. Taken together, these findings suggest that methyl sartortuoate is capable of leading to activation of caspase-8, -9, -3, increasing p53 and Bax/Bcl-2 ratio apoptosis through MAPK-dependent apoptosis and results in G2-M phase arrest in LoVo and RKO cells. Thus, methyl sartortuoate may be a promising anticancer candidate.

  3. Uranium induces apoptosis in lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Periyakaruppan, Adaikkappan; Sarkar, Shubhashish; Sadanandan, Bindu; Thomas, Renard; Wilson, Bobby L. [Texas Southern University, Environmental Toxicology Program, Department of Chemistry, Houston, TX (United States); Ravichandran, Prabakaran; Sharma, Chidananda S.; Ramesh, Vani; Hall, Joseph C.; Ramesh, Govindarajan T. [Norfolk State University, Molecular Toxicology Laboratory, Department of Biology, Center for Biotechnology and Biomedical Sciences, Norfolk, VA (United States)

    2009-06-15

    Uranium is a naturally occurring radioactive material present everywhere in the environment. It is toxic because of its chemical or radioactive properties. Uranium enters environment mainly from mines and industry and cause threat to human health by accumulating in lungs as a result of inhalation. In our previous study, we have shown the effectiveness of antioxidant system response to the oxidative stress induced by uranyl acetate (UA) in rat lung epithelial (LE) cells. As part of our continuing studies; here, we investigated the mechanism underlying when LE cells are exposed to different concentration of UA. Oxidative stress may lead to apoptotic signaling pathways. LE cells treated with 0.25, 0.5 and 1 mM of UA results in dose and time-dependent increase in activity of both caspases-3 and -8. Increase in the concentration of cytochrome-c oxidase in cytosol was seen in LE cells treated with 1 mM UA as a result of mitochondria membrane permeability. The cytochrome-c leakage may trigger the apoptotic pathway. TUNEL assay performed in LE cells treated with 1 mM of UA showed significant incorporation of dNTPs in the nucleus after 24 h. In the presence of the caspase inhibitors, we observed the significant decrease in the activity of caspases-8 and -3 in 0.5 and 1 mM UA-treated LE cells. (orig.)

  4. A New Diterpene from Litsea cubeba Fruits: Structure Elucidation and Capability to Induce Apoptosis in HeLa Cells

    Directory of Open Access Journals (Sweden)

    Piyapat Trisonthi

    2014-05-01

    Full Text Available A new diterpene, identified as (+-6-(4-hydroxy-4-methyl-2-pentenoyl-4,6-dimethyl-5-(3-methyl-2-butenyl-1,3-cyclohexadienecarbaldehyde (1, cubelin, was isolated from a methanol extract of Litsea cubeba fruits by normal phase column chromatography and purified by preparative HPLC. The structure elucidation was conducted by spectroscopic methods (UV, IR, ESI-TOF-MS, 1-D and 2-D NMR. Cubelin exhibited activity against HeLa cell viability and proliferation. The cells also exhibited changes in nuclear morphology which are hallmarks of apoptotic cell death. The presence of cleaved caspase-3/-7, caspase-8 and caspase-9 in the cubelin treated population indicated the potential of the compound to induce apoptosis in HeLa cells via both intrinsic and extrinsic pathways.

  5. TRAIL-Induced Caspase Activation Is a Prerequisite for Activation of the Endoplasmic Reticulum Stress-Induced Signal Transduction Pathways.

    Science.gov (United States)

    Lee, Dae-Hee; Sung, Ki Sa; Guo, Zong Sheng; Kwon, William Taehyung; Bartlett, David L; Oh, Sang Cheul; Kwon, Yong Tae; Lee, Yong J

    2016-05-01

    It is well known that tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis can be initially triggered by surface death receptors (the extrinsic pathway) and subsequently amplified through mitochondrial dysfunction (the intrinsic pathway). However, little is known about signaling pathways activated by the TRAIL-induced endoplasmic reticulum (ER) stress response. In this study, we report that TRAIL-induced apoptosis is associated with the endoplasmic reticulum (ER) stress response. Human colorectal carcinoma HCT116 cells were treated with TRAIL and the ER stress-induced signal transduction pathway was investigated. During TRAIL treatment, expression of ER stress marker genes, in particular the BiP (binding immunoglobulin protein) gene, was increased and activation of the PERK (PKR-like ER kinase)-eIF2α (eukaryotic initiation factor 2α)-ATF4 (activating transcription factor 4)-CHOP (CCAAT-enhancer-binding protein homologous protein) apoptotic signal transduction pathway occurred. Experimental data from use of a siRNA (small interfering RNA) technique, caspase inhibitor, and caspase-3-deficient cell line revealed that TRAIL-induced caspase activation is a prerequisite for the TRAIL-induced ER stress response. TRAIL-induced ER stress was triggered by caspase-8-mediated cleavage of BAP31 (B cell receptor-associated protein 31). The involvement of the proapoptotic PERK-CHOP pathway in TRAIL-induced apoptosis was verified by using a PERK knockout (PERK(-/-)) mouse embryo fibroblast (MEF) cell line and a CHOP(-/-) MEF cell line. These results suggest that TRAIL-induced the activation of ER stress response plays a role in TRAIL-induced apoptotic death.

  6. Electronic properties of hydrogen- and oxygen-terminated diamond surfaces exposed to the air

    Institute of Scientific and Technical Information of China (English)

    Liu Feng-Bin; Wang Jia-Dao; Chen Da-Rong; Yan Da-Yun

    2009-01-01

    The electronic properties of hydrogen- and oxygen-terminated diamond surfaces exposed to the air are investigated by scanning probe microscopy (SPM). The results indicate that for the hydrogen-terminated diamond surface a shallow acceptor above the valence-band-maximum (VBM) appears in the band gap. However, the oxygen-terminated diamond film exhibits a high resistivity with a wide band gap. Based on the density-functional-theory, the densities of states, corresponding to molecular adsorbate in hydrogenated and oxygenated diamond (100) surfaces, are studied. The results show that the shallow acceptor in the band gap for the hydrogen-terminated diamond film can be attributed to the interaction between the surface C-H bonding orbitals and the adsorbate molecules, while for the oxygen-terminated diamond film, the interaction between the surface C-O bonding orbitals and the adsorbate molecules can induce occupied states in the valence-band.

  7. Sanguinarine induces apoptosis of human osteosarcoma cells through the extrinsic and intrinsic pathways

    Energy Technology Data Exchange (ETDEWEB)

    Park, Hyunjin; Bergeron, Eric; Senta, Helena; Guillemette, Kim [Cell-Biomaterial Biohybrid Systems, Universite de Sherbrooke, Department of Chemical Engineering and Biotechnological Engineering, Canada J1K 2R1 (Canada); Beauvais, Sabrina [Cell-Biomaterial Biohybrid Systems, Universite de Sherbrooke, Department of Chemical Engineering and Biotechnological Engineering, Canada J1K 2R1 (Canada); Development of Bioprocess, Universite de Sherbrooke, Department of Chemical Engineering and Biotechnological Engineering, Canada J1K 2R1 (Canada); Blouin, Richard [Universite de Sherbrooke, Department of Biology, Canada J1K 2R1 (Canada); Sirois, Joel [Development of Bioprocess, Universite de Sherbrooke, Department of Chemical Engineering and Biotechnological Engineering, Canada J1K 2R1 (Canada); Faucheux, Nathalie, E-mail: Nathalie.Faucheux@Usherbrooke.ca [Cell-Biomaterial Biohybrid Systems, Universite de Sherbrooke, Department of Chemical Engineering and Biotechnological Engineering, Canada J1K 2R1 (Canada)

    2010-08-27

    Research highlights: {yields} We show for the first time the effect of sanguinarine (SA) on MG63 and SaOS-2 cells. {yields} SA altered osteosarcoma cell viability in a concentration and time dependent manner. {yields} SA induced osteosarcoma cell apoptosis and increased caspase-8 and -9 activities. {yields} SA decreased dose dependently the Bcl-2 protein level only in MG63 cells. {yields} SaOS-2 which are osteoblast-derived, seemed more resistant to SA than MG63. -- Abstract: The quaternary benzo[c]phenanthridine alkaloid sanguinarine inhibits the proliferation of cancerous cells from different origins, including lung, breast, pancreatic and colon, but nothing is known of its effects on osteosarcoma, a primary malignant bone tumour. We have found that sanguinarine alters the morphology and reduces the viability of MG-63 and SaOS-2 human osteosarcoma cell lines in concentration- and time-dependent manner. Incubation with 1 {mu}mol/L sanguinarine for 4 and 24 h killed more efficiently MG-63 cells than SaOS-2 cells, while incubation with 5 {mu}mol/L sanguinarine killed almost 100% of both cell populations within 24 h. This treatment also changed the mitochondrial membrane potential in both MG-63 and SaOS-2 cells within 1 h, caused chromatin condensation and the formation of apoptotic bodies. It activated multicaspases, and increased the activities of caspase-8 and caspase-9 in both MG-63 and SaOS-2 cells. These data highlight sanguinarine as a novel potential agent for bone cancer therapy.

  8. Effect of magnetic nanoparticles on apoptosis and cell cycle induced by wogonin in Raji cells

    Directory of Open Access Journals (Sweden)

    Wang XM

    2012-02-01

    Full Text Available Lei Wang1,2,*, Haijun Zhang1,2,*, Baoan Chen1,2, Guohua Xia1,2, Shuai Wang1,2, Jian Cheng1,2, Zeye Shao1,2, Chong Gao1,2, Wen Bao1,2, Liang Tian1,2, Yanyan Ren1,2, Peipei Xu1,2, Xiaohui Cai1,2, Ran Liu1,2, Xuemei Wang3 1Department of Hematology and Oncology, Zhongda Hospital, Medical School, 2Faculty of Oncology, Medical School, 3State Key Laboratory of Bioelectronics (Chien-Shiung Wu Laboratory, Southeast University, Nanjing, China*These authors contributed equally to this workAbstract: Traditional Chinese medicine is gradually becoming a new source of anticancer drugs. One such example is wogonin, which is cytotoxic to various cancer cell lines in vitro. However, due to its low water solubility, wogonin is restricted to clinical administration. Recently, the application of drug-coated magnetic nanoparticles (MNPs to increase water solubility of the drug and to enhance its chemotherapeutic efficiency has attracted much attention. In this study, wogonin was conjugated with the drug delivery system of MNPs by mechanical absorption polymerization to fabricate wogonin-loaded MNPs. It was demonstrated that MNPs could strengthen wogonin-induced cell inhibition, apoptosis, and cell cycle arrest in Raji cells by methylthiazol tetrazolium assay, flow cytometer assay, and nuclear 4',6-diamidino-2-phenylindole staining. Furthermore, the molecular mechanisms of these phenomena were explored by western blot, in which the protein levels of caspase 8 and caspase 3 were increased significantly while those of survivin and cyclin E were decreased significantly in wogonin-MNPs group. These findings suggest that the combination of wogonin and MNPs provides a promising strategy for lymphoma therapy.Keywords: wogonin, magnetic nanoparticles, Raji cell, apoptosis, cell cycle, caspase 8, caspase 3, survivin, cyclin E

  9. Increased apoptosis of lactotrophs in streptozotocin-induced diabetic rats is followed by increased proliferation.

    Science.gov (United States)

    Arroba, Ana I; Lechuga-Sancho, Alfonso M; Frago, Laura M; Argente, Jesús; Chowen, Julie A

    2006-10-01

    Poorly controlled diabetes mellitus can result in decreased prolactin production and thus problems with lactation, reproduction, and other physiological processes. This may be due to a loss of lactotrophs, as we have previously shown that long-term (8 weeks) poorly controlled streptozotocin-induced diabetes results in increased death of lactotrophs and that this most likely occurs through the activation of caspase-8 and the extrinsic cell death cascade. However, cell proliferation is also increased in the anterior pituitary at this time, although the cell type undergoing this proliferation and whether it is a response to the increased cell death remains unknown. In order to determine the time-course of increased cell death and proliferation in the anterior pituitary and if this is related to changes in tumor necrosis factor (TNF)-alpha, a cytokine involved in the activation of the extrinsic cell death pathway, rats were killed at 1, 4, 6, and 8 weeks after the induction of diabetes. Cell death was significantly increased after 4 weeks, as was caspase-8 activation, although circulating levels of TNF-alpha were increased as early as 1 week. Pituitary levels of TNF-alpha did not change significantly until 8 weeks after diabetes onset. Similarly, Western-blot analysis of proliferating cell nuclear antigen showed that anterior pituitary cell proliferation increased significantly 8 weeks after diabetes onset, with the majority of proliferating cells, as detected by BrdU incorporation, corresponding to lactotrophs. These results suggest that the increased death of lactotrophs in poorly controlled diabetic rats is followed by increased proliferation of this cell type, even when no treatment is given.

  10. Gambling with video lottery terminals.

    Science.gov (United States)

    Doiron, J P; Mazer, D B

    2001-09-01

    Semistructured interviews were conducted with 7 persons who had significant involvements with video lottery terminal (VLT) gambling, and themes associated with different phases of the gambling experience were identified. The preinvolvement phase was characterized by lack of meaningful relationships, problematic relationships, and feelings of loss. Early involvement reflected attempts to "fill the void" and the casual innocence of initial VLT playing. The deepening involvement phase indicated themes of the language of relationship, for example, focused engagement, emotional highs and lows, and the escape and competition offered by gambling. Ending involvement themes included the emotional difficulty of quitting and strategies used to break the habit. Implications of these results for models of addiction and for the treatment of gambling problems are explored.

  11. Apocynin attenuates cholesterol oxidation product-induced programmed cell death by suppressing NF-κB-mediated cell death process in differentiated PC12 cells.

    Science.gov (United States)

    Lee, Da Hee; Nam, Yoon Jeong; Lee, Chung Soo

    2015-10-01

    Cholesterol oxidation products are suggested to be involved in neuronal degeneration. Apocynin has demonstrated to have anti-inflammatory and anti-oxidant effects. We assessed the effect of apocynin on the cholesterol oxidation product-induced programmed cell death in neuronal cells using differentiated PC12 cells in relation to NF-κB-mediated cell death process. 7-Ketocholesterol and 25-hydroxycholesterol decreased the levels of Bid and Bcl-2, increased the levels of Bax and p53, and induced loss of the mitochondrial transmembrane potential, release of cytochrome c and activation of caspases (-8, -9 and -3). 7-Ketocholesterol caused an increase in the levels of cytosolic and nuclear NF-κB p65, cytosolic NF-κB p50 and cytosolic phospho-IκB-α, which was inhibited by the addition of 0.5 μM Bay11-7085 (an inhibitor of NF-κB activation). Apocynin attenuated the cholesterol oxidation product-induced changes in the programmed cell death-related protein levels, NF-κB activation, production of reactive oxygen species, and depletion of GSH. The results show that apocynin appears to attenuate the cholesterol oxidation product-induced programmed cell death in PC12 cells by suppressing the activation of the mitochondrial pathway and the caspase-8- and Bid-dependent pathways that are mediated by NF-κB activation. The preventive effect appears to be associated with the inhibitory effect on the production of reactive oxygen species and depletion of GSH.

  12. Sulforaphane-induced apoptosis in human leukemia HL-60 cells through extrinsic and intrinsic signal pathways and altering associated genes expression assayed by cDNA microarray.

    Science.gov (United States)

    Shang, Hung-Sheng; Shih, Yung-Luen; Lee, Ching-Hsiao; Hsueh, Shu-Ching; Liu, Jia-You; Liao, Nien-Chieh; Chen, Yung-Liang; Huang, Yi-Ping; Lu, Hsu-Feng; Chung, Jing-Gung

    2017-01-01

    Sulforaphane (SFN), one of the isothiocyanates, is a biologically active compound extracted from cruciferous vegetables, and has been shown to induce cytotoxic effects on many human cancer cells including human leukemia cells. However, the exact molecular mechanism and altered gene expression associated with apoptosis is unclear. In this study, we investigated SFN-induced cytotoxic effects and whether or not they went through cell-cycle arrest and induction of apoptosis and further examined molecular mechanism and altered gene expression in human leukemia HL-60 cells. Cell viability, cell-cycle distribution, sub-G1 (apoptosis), reactive oxygen species (ROS) and Ca(2+) production, levels of mitochondrial membrane potential (ΔΨm ), and caspase-3, -8, and -9 activities were assayed by flow cytometry. Apoptosis-associated proteins levels and gene expressions were examined by Western blotting and cDNA microarray assays, respectively. Results indicated that SFN decreased viable cells, induced G2/M phase arrest and apoptosis based on sub-G1 phase development. Furthermore, SFN increased ROS and Ca(2+) production and decreased the levels of ΔΨm and activated caspase-3, -8, and -9 activities in HL-60 cells. SFN significantly upregulated the expression of BAX, Bid, Fas, Fas-L, caspase-8, Endo G, AIF, and cytochrome c, and inhibited the antiapoptotic proteins such as Bcl-x and XIAP, that is associated with apoptosis. We also used cDNA microarray to confirm several gene expressions such as caspase -8, -3, -4, -6, and -7 that are affected by SFN. Those results indicated that SFN induced apoptosis in HL-60 cells via Fas- and mitochondria-dependent pathways. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 311-328, 2017.

  13. The selective protein kinase C inhibitor, Ro-31-8220, inhibits mitogen-activated protein kinase phosphatase-1 (MKP-1) expression, induces c-Jun expression, and activates Jun N-terminal kinase.

    Science.gov (United States)

    Beltman, J; McCormick, F; Cook, S J

    1996-10-25

    The role of protein kinase C (PKC) in inflammation, mitogenesis, and differentiation has been deduced in part through the use of a variety of PKC inhibitors. Two widely used inhibitors are the structurally related compounds GF109203X and Ro-31-8220, both of which potently inhibit PKC activity and are believed to be highly selective. While using GF109203X and Ro-31-8220 to address the role of PKC in immediate early gene expression, we observed striking differential effects by each of these two compounds. Growth factors induce the expression of the immediate early gene products MAP kinase phosphatase-1 (MKP-1), c-Fos and c-Jun. Ro-31-8220 inhibits growth factor-stimulated expression of MKP-1 and c-Fos but strongly stimulated c-Jun expression, even in the absence of growth factors. GF109203X displays none of these properties. These data suggest that Ro-31-8220 may have other pharmacological actions in addition to PKC inhibition. Indeed, Ro-31-8220 strongly stimulates the stress-activated protein kinase, JNK1. Furthermore, Ro-31-8220 apparently activates JNK in a PKC-independent manner. Neither the down-regulation of PKC by phorbol esters nor the inhibition of PKC by GF109203X affected the ability of Ro-31-8220 to activate JNK1. These data suggest that, in addition to potently inhibiting PKC, Ro-31-8220 exhibits novel pharmacological properties which are independent of its ability to inhibit PKC.

  14. Rac-1 superactivation triggers insulin-independent glucose transporter 4 (GLUT4) translocation that bypasses signaling defects exerted by c-Jun N-terminal kinase (JNK)- and ceramide-induced insulin resistance.

    Science.gov (United States)

    Chiu, Tim Ting; Sun, Yi; Koshkina, Alexandra; Klip, Amira

    2013-06-14

    Insulin activates a cascade of signaling molecules, including Rac-1, Akt, and AS160, to promote the net gain of glucose transporter 4 (GLUT4) at the plasma membrane of muscle cells. Interestingly, constitutively active Rac-1 expression results in a hormone-independent increase in surface GLUT4; however, the molecular mechanism and significance behind this effect remain unresolved. Using L6 myoblasts stably expressing myc-tagged GLUT4, we found that overexpression of constitutively active but not wild-type Rac-1 sufficed to drive GLUT4 translocation to the membrane of comparable magnitude with that elicited by insulin. Stimulation of endogenous Rac-1 by Tiam1 overexpression elicited a similar hormone-independent gain in surface GLUT4. This effect on GLUT4 traffic could also be reproduced by acutely activating a Rac-1 construct via rapamycin-mediated heterodimerization. Strategies triggering Rac-1 "superactivation" (i.e. to levels above those attained by insulin alone) produced a modest gain in plasma membrane phosphatidylinositol 3,4,5-trisphosphate, moderate Akt activation, and substantial AS160 phosphorylation, which translated into GLUT4 translocation and negated the requirement for IRS-1. This unique signaling capacity exerted by Rac-1 superactivation bypassed the defects imposed by JNK- and ceramide-induced insulin resistance and allowed full and partial restoration of the GLUT4 translocation response, respectively. We propose that potent elevation of Rac-1 activation alone suffices to drive insulin-independent GLUT4 translocation in muscle cells, and such a strategy might be exploited to bypass signaling defects during insulin resistance.

  15. Termination of floating-point computations

    OpenAIRE

    Serebrenik, Alexander; De Schreye, Danny

    2005-01-01

    Numerical computations form an essential part of almost any real-world program. Traditional approaches to termination of logic programs are restricted to domains isomorphic to (N,>); more recent works study termination of integer computations where the lack of well-foundedness of the integers has to be taken into account. Termination of computations involving floating-point numbers can be counterintuitive because of rounding errors and implementation conventions. We present a novel technique ...

  16. Ursodeoxycholic Acid Induces Death Receptor-mediated Apoptosis in Prostate Cancer Cells

    Science.gov (United States)

    Lee, Won Sup; Jung, Ji Hyun; Panchanathan, Radha; Yun, Jeong Won; Kim, Dong Hoon; Kim, Hye Jung; Kim, Gon Sup; Ryu, Chung Ho; Shin, Sung Chul; Hong, Soon Chan; Choi, Yung Hyun; Jung, Jin-Myung

    2017-01-01

    Background Bile acids have anti-cancer properties in a certain types of cancers. We determined anticancer activity and its underlying molecular mechanism of ursodeoxycholic acid (UDCA) in human DU145 prostate cancer cells. Methods Cell viability was measured with an MTT assay. UDCA-induced apoptosis was determined with flow cytometric analysis. The expression levels of apoptosis-related signaling proteins were examined with Western blotting. Results UDCA treatment significantly inhibited cell growth of DU145 in a dose-dependent manner. It induced cellular shrinkage and cytoplasmic blebs and accumulated the cells with sub-G1 DNA contents. Moreover, UDCA activated caspase 8, suggesting that UDCA-induced apoptosis is associated with extrinsic pathway. Consistent to this finding, UDCA increased the expressions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, death receptor 4 (DR4) and death receptor 5 (DR5), and TRAIL augmented the UDCA-induced cell death in DU145 cells. In addition, UDCA also increased the expressions of Bax and cytochrome c and decreased the expression of Bcl-xL in DU145 cells. This finding suggests that UDCA-induced apoptosis may be involved in intrinsic pathway. Conclusions UDCA induces apoptosis via extrinsic pathway as well as intrinsic pathway in DU145 prostate cancer cells. UDCA may be a promising anti-cancer agent against prostate cancer.

  17. Mitochondrial control of cell death induced by hyperosmotic stress.

    Science.gov (United States)

    Criollo, Alfredo; Galluzzi, Lorenzo; Maiuri, M Chiara; Tasdemir, Ezgi; Lavandero, Sergio; Kroemer, Guido

    2007-01-01

    HeLa and HCT116 cells respond differentially to sorbitol, an osmolyte able to induce hypertonic stress. In these models, sorbitol promoted the phenotypic manifestations of early apoptosis followed by complete loss of viability in a time-, dose-, and cell type-specific fashion, by eliciting distinct yet partially overlapping molecular pathways. In HCT116 but not in HeLa cells, sorbitol caused the mitochondrial release of the caspase-independent death effector AIF, whereas in both cell lines cytochrome c was retained in mitochondria. Despite cytochrome c retention, HeLa cells exhibited the progressive activation of caspase-3, presumably due to the prior activation of caspase-8. Accordingly, caspase inhibition prevented sorbitol-induced killing in HeLa, but only partially in HCT116 cells. Both the knock-out of Bax in HCT116 cells and the knock-down of Bax in A549 cells by RNA interference reduced the AIF release and/or the mitochondrial alterations. While the knock-down of Bcl-2/Bcl-X(L) sensitized to sorbitol-induced killing, overexpression of a Bcl-2 variant that specifically localizes to mitochondria (but not of the wild-type nor of a endoplasmic reticulum-targeted form) strongly inhibited sorbitol effects. Thus, hyperosmotic stress kills cells by triggering different molecular pathways, which converge at mitochondria where pro- and anti-apoptotic members of the Bcl-2 family exert their control.

  18. Multiterminal Conductance and Decoherence Effect of a Three-Terminal Kondo Dot

    Institute of Scientific and Technical Information of China (English)

    FANG Tie-Feng; WANG Shun-Jin

    2006-01-01

    @@ A three-terminal Kondo dot modelled by the Anderson Hamiltonian is investigated. In the strong correlation limit, we calculate the multiterminal conductance and the voltage-induced characteristic splitting of the nonequilibrium Kondo resonance by using the equation of motion approach from viewpoint of the correlation dynamics.A qualitative and reasonable agreement with a recently reported experiment is obtained. We also simulate phenomenologically the decoherence of the Kondo-coherent state formed in the two-terminal setup in the framework of our three-terminal model.

  19. Cpeb4-mediated translational regulatory circuitry controls terminal erythroid differentiation.

    Science.gov (United States)

    Hu, Wenqian; Yuan, Bingbing; Lodish, Harvey F

    2014-09-29

    While we have considerable understanding of the transcriptional networks controlling mammalian cell differentiation, our knowledge of posttranscriptional regulatory events is very limited. Using differentiation of primary erythroid cells as a model, we show that the sequence-specific mRNA-binding protein Cpeb4 is strongly induced by the erythroid-important transcription factors Gata1 and Tal1 and is essential for terminal erythropoiesis. By interacting with the translation initiation factor eIF3, Cpeb4 represses the translation of a large set of mRNAs, including its own mRNA. Thus, transcriptional induction and translational repression combine to form a negative feedback loop to control Cpeb4 protein levels within a specific range that is required for terminal erythropoiesis. Our study provides an example of how translational control is integrated with transcriptional regulation to precisely control gene expression during mammalian cell differentiation.

  20. Neutral atom transport from the termination shock to 1 AU

    CERN Document Server

    Bzowski, M; Bzowski, Maciej; Tarnopolski, Slawomir

    2006-01-01

    Dynamics of H, D, and heavy Energetic Neutral Atoms (ENA) between the termination shock and 1 AU is discussed in the context of the forthcoming NASA SMEX mission IBEX. In particular, effects of the velocity-dependent radiation pressure on atomic trajectories are considered and ionization losses between TS and 1 AU are studied. It is shown, among others, that most of the dynamical effects and ionization losses are induced within a few AU from the Sun, which translates to the time domain into $\\sim 1 - 3$ solar rotations before detection. This loosens considerably time requirements for tracking the ionization and radiation pressure history to just prior 3 months. ENA seem excellent tracers of the processes within the heliospheric interface, with the transport effects between the termination shock and detector relatively mild and easy to account for.

  1. St. James marine terminal facility description

    Energy Technology Data Exchange (ETDEWEB)

    1993-10-01

    The US Department of Energy (DOE) currently owns and operates a marine terminal on the west bank of the Mississippi River at St. James, Louisiana. The St. James facility was constructed by the Department to provide marine services associated with the fill and drawdown of the Strategic Petroleum Reserve (SPR) crude oil storage facilities located at Bayou Choctaw and Weeks Island, Louisiana. Although strategic to the mission of the SPR in the event of a national emergency, the St. James terminal is situated such that it has a high potential to also serve the commercial industry`s needs for crude oil terminalling and storage. The St. James terminal is located approximately 45 miles west of New Orleans and 30 miles southeast of Baton Rouge, and approximately 160 miles upstream from the mouth of the Mississippi River. Construction of the St. James terminal was initiated in 1978 and was completed in 1980. Since then, the terminal has received and transferred over 125 million barrels of crude oil to the SPR sites for storage. For crude oil distribution, the St. James terminal was connected to the neighboring LOCAP terminal by a 0.1 mile 36-inch pipeline in 1981 and to the Capline terminal by a 0.5 mile 30-inch pipeline in 1988. The terminal also has a 30-inch pipeline connection to the Koch oil terminal which was used for initial fill purposes; however, this pipeline has been disconnected and is currently inactive. A complete description of the St. James terminal facilities, operational capabilities, operational certifications, and future Government requirements are presented in Sections 2, 3, 4, and 5 respectively.

  2. 77 FR 38128 - Withdrawal of TORP Terminal LP, Bienville Offshore Energy Terminal Liquefied Natural Gas (LNG...

    Science.gov (United States)

    2012-06-26

    ... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF TRANSPORTATION Maritime Administration Withdrawal of TORP Terminal LP, Bienville Offshore Energy Terminal Liquefied Natural Gas (LNG) Deepwater Port Application AGENCY: Maritime Administration, DOT. ACTION: Deepwater...

  3. Termination review committees: are they necessary?

    Science.gov (United States)

    Woodrow, Nicole L

    2003-07-21

    In Victoria, decisions regarding late termination of pregnancy no longer involve just pregnant women and their clinicians. At two major women's hospitals, committees now govern the decision-making process for approval of a late termination of pregnancy. The legal and ethical implications of clinical decision-making by committee need to be widely debated.

  4. Ligand sphere conversions in terminal carbide complexes

    DEFF Research Database (Denmark)

    Morsing, Thorbjørn Juul; Reinholdt, Anders; Sauer, Stephan P. A.

    2016-01-01

    Metathesis is introduced as a preparative route to terminal carbide complexes. The chloride ligands of the terminal carbide complex [RuC(Cl)2(PCy3)2] (RuC) can be exchanged, paving the way for a systematic variation of the ligand sphere. A series of substituted complexes, including the first exam...

  5. The OCLC Terminal: An Instruction Manual.

    Science.gov (United States)

    Arnold, Jane; And Others

    Prepared for users of OCLC terminals at the Michigan State University library, this manual provides a brief description of the contents of the OCLC database, instructions for constructing search requests in OCLC, guidelines for interpreting OCLC search results, a key to "reading" the OCLC screen, an overview of typical terminal responses…

  6. [Terminal phase hydration, pain and delirium

    DEFF Research Database (Denmark)

    Heick, A.

    2009-01-01

    Hydration of the terminal patient may relieve confusion and complaints of "dry mouth". But it may worsen oedema of the brain, lungs, and extremities, worsen terminal rattling and cause a need for frequent changing of diapers. The decision of whether and how to treat a dying patient with fluids...

  7. Size-change Termination and Bound Analysis

    DEFF Research Database (Denmark)

    Avery, James Emil

    2006-01-01

    Despite its simplicity, the size-change termination principle, presented by Lee, Jones and Ben-Amram in [LJB01], is surprisingly strong and is able to show termination for a large class of programs. A significant limitation for its use, however, is the fact that the SCT requires data types...

  8. 77 FR 13322 - Termination of Dormant Proceedings

    Science.gov (United States)

    2012-03-06

    ... COMMISSION Termination of Dormant Proceedings AGENCY: Federal Communications Commission. ACTION: Notice... comment on whether certain docketed Commission proceedings should be terminated as dormant. The Commission... Proceedings as Dormant, document DA 12-220, released on February 15, 2012 in CG Docket No. 12-39. The...

  9. Expression of survivin and p53 modulates honokiol-induced apoptosis in colorectal cancer cells.

    Science.gov (United States)

    Lai, Ying-Jiun; Lin, Chien-I; Wang, Chia-Lin; Chao, Jui-I

    2014-11-01

    Honokiol is a small biphenolic compound, which exerts antitumor activities; however, the precise mechanism of honokiol-induced apoptosis in the human colorectal cancer cells remains unclear. Here, we show that survivin and p53 display the opposite role on the regulation of honokiol-induced apoptosis in the human colorectal cancer cells. Honokiol induced the cell death and apoptosis in various colorectal cancer cell lines. Moreover, honokiol elicited the extrinsic death receptor pathway of DR5 and caspase 8 and the intrinsic pathway of caspase 9. The common intrinsic and extrinsic downstream targets of activated caspase 3 and PARP protein cleavage were induced by honokiol. Interestingly, honokiol reduced anti-apoptotic survivin protein and gene expression. Transfection with a green fluorescent protein (GFP)-survivin-expressed vector increased the colorectal cancer cell viability and resisted the honokiol-induced apoptosis. Meantime, honokiol increased total p53 and the phosphorylated p53 proteins at Ser15 and Ser46. The p53-wild type colorectal cancer cells were exhibited greater cytotoxicity, apoptosis and survivin reduction than the p53-null cancer cells after treatment with honokiol. Together, these findings demonstrate that the existence of survivin and p53 can modulate the honokiol-induced apoptosis in the human colorectal cancer cells.

  10. 78 FR 5717 - Safety Zone; Military Ocean Terminal Concord Safety Zone, Suisun Bay, Military Ocean Terminal...

    Science.gov (United States)

    2013-01-28

    ... SECURITY Coast Guard 33 CFR Part 165 RIN 1625-AA00 Safety Zone; Military Ocean Terminal Concord Safety Zone, Suisun Bay, Military Ocean Terminal Concord, CA AGENCY: Coast Guard, DHS. ACTION: Interim rule and... Suisun Bay near Military Ocean Terminal Concord, CA in support of military onload and offload...

  11. Mood and audience effects on video lottery terminal gambling.

    Science.gov (United States)

    Mishra, Sandeep; Morgan, Michael; Lalumière, Martin L; Williams, Robert J

    2010-09-01

    Little is known about the situational factors associated with gambling behavior. We induced 180 male participants (mean age: 21.6) into a positive, negative, or neutral mood prior to gambling on a video lottery terminal (VLT). While gambling, participants were observed by either a male peer, female peer, or no one. Induced mood had no effect on gambling behavior. Participants induced into a negative mood prior to gambling, however, reported more positive moods after gambling, whereas those with positive and neutral moods reported more negative moods after gambling. Participants observed by either a male or female peer spent less time gambling on the VLT compared to those not observed. Participants observed by a female peer lost less money relative to the other observer conditions. Degree of problem gambling in the last year had little influence on these effects. Some practical implications of these findings are discussed.

  12. An Easy-To-Use Combination Four-Terminal-Pair/Two-Terminal-Pair AC Transformer Bridge.

    Science.gov (United States)

    Jeffery, A; Shields, J Q; Lee, L H

    1998-01-01

    A new four-terminal-pair bridge, capable of achieving a relative standard uncertainty of 1×10(-9), was constructed at the National Institute of Standards and Technology by converting a two-terminal-pair bridge. The conversion requires only the addition of components which are easily removed if two-terminal-pair measurements are to be made. The design and testing of this bridge is described. The new four-terminal-pair bridge requires fewer auxiliary balances than the present four-terminal-pair bridge employed at NIST, which makes it much easier to use. This new design can be used to compare capacitance, resistance, and inductance standards.

  13. Chestnut extract induces apoptosis in AGS human gastric cancer cells.

    Science.gov (United States)

    Lee, Hyun Sook; Kim, Eun Ji; Kim, Sun Hyo

    2011-06-01

    In Korea, chestnut production is increasing each year, but consumption is far below production. We investigated the effect of chestnut extracts on antioxidant activity and anticancer effects. Ethanol extracts of raw chestnut (RCE) or chestnut powder (CPE) had dose-dependent superoxide scavenging activity. Viable numbers of MDA-MD-231 human breast cancer cells, DU145 human prostate cancer cells, and AGS human gastric cancer cells decreased by 18, 31, and 69%, respectively, following treatment with 200 µg/mL CPE for 24 hr. CPE at various concentrations (0-200 µg/mL) markedly decreased AGS cell viability and increased apoptotic cell death dose and time dependently. CPE increased the levels of cleaved caspase-8, -7, -3, and poly (ADP-ribose) polymerase in a dose-dependent manner but not cleaved caspase-9. CPE exerted no effects on Bcl-2 and Bax levels. The level of X-linked inhibitor of apoptosis protein decreased within a narrow range following CPE treatment. The levels of Trail, DR4, and Fas-L increased dose-dependently in CPE-treated AGS cells. These results show that CPE decreases growth and induces apoptosis in AGS gastric cancer cells and that activation of the death receptor pathway contributes to CPE-induced apoptosis in AGS cells. In conclusion, CPE had more of an effect on gastric cancer cells than breast or prostate cancer cells, suggesting that chestnuts would have a positive effect against gastric cancer.

  14. Participation of Private Investors in Container Terminal Operation: Influence of Global Terminal Operators

    Directory of Open Access Journals (Sweden)

    Heejung YEO

    2015-09-01

    Full Text Available Private container terminal operators have begun to participate in the port business in Asia since the late 1980s. Terminal operators decide whether to invest in terminal infrastructures, and the enhancement of the quality of service level. The paper analyses the influence of global terminal operators and the port ownership structure on the container terminal's efficiency. Two hundred and sixty container terminal data for China, Korea, and Japan were collected. The paper applies a negative binomial regression analysis. The paper finds that the port restructuring has contributed to productivity gains. It is found that the influence of GTO on the efficiency is evident and positively related to the port efficiency. The paper also finds that the country effect prevails over a terminal operator group effect.

  15. Traditional Herbal Medicine, Rikkunshito, Induces HSP60 and Enhances Cytoprotection of Small Intestinal Mucosal Cells as a Nontoxic Chaperone Inducer

    Directory of Open Access Journals (Sweden)

    Kumiko Tamaki

    2012-01-01

    Full Text Available Increasing incidence of small intestinal ulcers associated with nonsteroidal anti-inflammatory drugs (NSAIDs has become a topic with recent advances of endoscopic technology. However, the pathogenesis and therapy are not fully understood. The aim of this study is to examine the effect of Rikkunshito (TJ-43, a traditional herbal medicine, on expression of HSP60 and cytoprotective ability in small intestinal cell line (IEC-6. Effect of TJ-43 on HSP60 expression in IEC-6 cells was evaluated by immunoblot analysis. The effect of TJ-43 on cytoprotective abilities of IEC-6 cells against hydrogen peroxide or indomethacin was studied by MTT assay, LDH-release assay, caspase-8 activity, and TUNEL. HSP60 was significantly induced by TJ-43. Cell necrosis and apoptosis were significantly suppressed in IEC-6 cells pretreated by TJ-43 with overexpression of HSP60. Our results suggested that HSP60 induced by TJ-43 might play an important role in protecting small intestinal epithelial cells from apoptosis and necrosis in vitro.

  16. 42 CFR 489.53 - Termination by CMS.

    Science.gov (United States)

    2010-10-01

    ... 42 Public Health 5 2010-10-01 2010-10-01 false Termination by CMS. 489.53 Section 489.53 Public... Reinstatement After Termination § 489.53 Termination by CMS. (a) Basis for termination of agreement with any provider. CMS may terminate the agreement with any provider if CMS finds that any of the following...

  17. A corrugated termination shock in pulsar wind nebulae?

    Science.gov (United States)

    Lemoine, Martin

    2016-08-01

    Successful phenomenological models of pulsar wind nebulae assume efficient dissipation of the Poynting flux of the magnetized electron-positron wind as well as efficient acceleration of the pairs in the vicinity of the termination shock, but how this is realized is not yet well understood. This paper suggests that the corrugation of the termination shock, at the onset of nonlinearity, may lead towards the desired phenomenology. Nonlinear corrugation of the termination shock would convert a fraction of order unity of the incoming ordered magnetic field into downstream turbulence, slowing down the flow to sub-relativistic velocities. The dissipation of turbulence would further preheat the pair population on short length scales, close to equipartition with the magnetic field, thereby reducing the initial high magnetization to values of order unity. Furthermore, it is speculated that the turbulence generated by the corrugation pattern may sustain a relativistic Fermi process, accelerating particles close to the radiation reaction limit, as observed in the Crab nebula. The required corrugation could be induced by the fast magnetosonic modes of downstream nebular turbulence; but it could also be produced by upstream turbulence, either carried by the wind or seeded in the precursor by the accelerated particles themselves.

  18. Ginsenoside Rh2 Showing Ability to Induce Apoptosis in HeLa Cells

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    This paper deals with the inhibitory mechanisms of ginsenoside G-Rh2 on the growth of tumor cells. G-Rh2 significantly inhibited the proliferation of human cervical adenocarcinoma HeLa cells in a time- and dose-dependent manner. G-Rh2 induced apoptotic manifestations in HeLa cells as evidenced by the changes in the cell morphology, the DNA fragmentation and the activation of caspases. Caspase inhibitors, caspase family inhibitor, z-Val-Ala-Asp-fmk(z-VAD-fmk); caspase-1 inhibitor, Ac-Tyr-Val-Ala-Asp-chloromethyl-ketone(Ac-YVAD-cmk); caspase-3 inhibitor, z-Asp-Glu-Val-Asp-fmk(z-DEVE-fmk) and caspase-8 inhibitor, z-Ile-Glu-Asp-fmk(z-IETD-fmk) effectively attenuated G-Rh2-induced cell death. The activities of caspase-1 and caspase-3 were increased in the G-Rh2-induced apoptotic process. However, caspase inhibitors can not inhibit G-Rh-2 induced cell death completely. These results suggest that G-Rh2-induced cell death is mediated by the activation of caspase cascade, but there might be some other pathways for induction of this apoptosis.

  19. Gliotoxin Isolated from Marine Fungus Aspergillus sp. Induces Apoptosis of Human Cervical Cancer and Chondrosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Van-Tinh Nguyen

    2013-12-01

    Full Text Available Gliotoxin, a secondary metabolite produced by marine fungus Aspergillus sp., possesses various biological activities including anticancer activity. However, the mechanism underlying gliotoxin-induced cytotoxicity on human cervical cancer (Hela and human chondrosarcoma (SW1353 cells remains unclear. In this study, we focused on the effect of gliotoxin induction on apoptosis, the activating expressions of caspase family enzymes in the cells. Apoptotic cell levels were measured through DAPI and Annexin V/Propidium Iodide (PI double staining analysis. The apoptotic protein expression of Bcl-2 and caspase family was detected by Western blot in Hela and SW1353 cells. Our results showed that gliotoxin treatment inhibited cell proliferation and induced significant morphological changes. Gliotoxin induced apoptosis was further confirmed by DNA fragmentation, chromatin condensation and disrupted mitochondrial membrane potential. Gliotoxin-induced activation of caspase-3, caspase-8 and caspase-9, down-regulation of Bcl-2, up-regulation of Bax and cytochromec (cyt c release showed evidence for the gliotoxin activity on apoptosis. These findings suggest that gliotoxin isolated from marine fungus Aspergillus sp. induced apoptosis in Hela and SW1353 cells via the mitochondrial pathway followed by downstream events leading to apoptotic mode of cell death.

  20. Operational Optimization in Port Container Terminals

    DEFF Research Database (Denmark)

    As a result of the significant increase in worldwide containerized transportation the development of efficient handling systems in marine terminals has become very important for port competitiveness. In order to optimize the productivity the total handling time for containers in the terminal must...... be minimized. An overview of the different operational problems in port container terminals is presented and an aggregated model and solution approach is shown. Next, there will be focused on the yard storage problem and a mathematical formulation and solution proposals will be presented....