Targeting apoptosis signaling in pancreatic cancer

The ability to escape apoptosis or programmed cell death is a hallmark of human cancers, for example pancreatic cancer. This can promote tumorigenesis, since too little cell death by apoptosis disturbs tissue homeostasis. Additionally, defective apoptosis signaling is the underlying cause of failure to respond to current treatment approaches, since therapy-mediated antitumor activity requires the intactness of apoptosis signaling pathways in cancer cells. Thus, the elucidation of defects in the regulation of apoptosis in pancreatic carcinoma can result in the identification of novel targets for therapeutic interference and for exploitation for cancer drug discovery. Keywords: apoptosis; pancreatic cancer; TRAIL; IAPs; mitochondri

Caspase-1 engagement and TLR-induced c-FLIP expression suppress ASC/caspase-8-dependent apoptosis by inflammasome sensors NLRP1b and NLRC4

The caspase activation and recruitment domain (CARD)-based inflammasome sensors NLRP1b and NLRC4 induce caspase-1-dependent pyroptosis independent of the inflammasome adaptor ASC. Here, we show that NLRP1b and NLRC4 trigger caspase-8-mediated apoptosis as an alternative cell death program in caspase-1(-/-) macrophages and intestinal epithelial organoids (IECs). The caspase-8 adaptor FADD was recruited to ASC specks, which served as cytosolic platforms for caspase-8 activation and NLRP1b/NLRC4-induced apoptosis. We further found that caspase-1 protease activity dominated over scaffolding functions in suppressing caspase-8 activation and induction of apoptosis of macrophages and IECs. Moreover, TLR-induced c-FLIP expression inhibited caspase-8-mediated apoptosis downstream of ASC speck assembly, but did not affect pyroptosis induction by NLRP1b and NLRC4. Moreover, unlike during pyroptosis, NLRP1b- and NLRC4-elicited apoptosis retained alarmins and the inflammasome-matured cytokines interleukin 1 beta (IL-1 beta) and IL-18 intracellularly. This work identifies critical mechanisms regulating apoptosis induction by the inflammasome sensors NLRP1b and NLRC4 and suggests converting pyroptosis into apoptosis as a paradigm for suppressing inflammation

Investigating the evolution of apoptosis in malaria parasites: the importance of ecology

Apoptosis is a precisely regulated process of cell death which occurs widely in multicellular organisms and is essential for normal development and immune defences. In recent years, interest has grown in the occurrence of apoptosis in unicellular organisms. In particular, as apoptosis has been reported in a wide range of species, including protozoan malaria parasites and trypanosomes, it may provide a novel target for intervention. However, it is important to understand when and why parasites employ an apoptosis strategy before the likely long-and short-term success of such an intervention can be evaluated. The occurrence of apoptosis in unicellular parasites provides a challenge for evolutionary theory to explain as organisms are expected to have evolved to maximise their own proliferation, not death. One possible explanation is that protozoan parasites undergo apoptosis in order to gain a group benefit from controlling their density as this prevents premature vector mortality. However, experimental manipulations to examine the ultimate causes behind apoptosis in parasites are lacking. In this review, we focus on malaria parasites to outline how an evolutionary framework can help make predictions about the ecological circumstances under which apoptosis could evolve. We then highlight the ecological considerations that should be taken into account when designing evolutionary experiments involving markers of cell death, and we call for collaboration between researchers in different fields to identify and develop appropriate markers in reference to parasite ecology and to resolve debates on terminology.Host-parasite interactio

A cardinal role for cathepsin D in co-ordinating the host-mediated apoptosis of macrophages and killing of pneumococci

The bactericidal function of macrophages against pneumococci is enhanced by their apoptotic demise, which is controlled by the anti-apoptotic protein Mcl-1. Here, we show that lysosomal membrane permeabilization (LMP) and cytosolic translocation of activated cathepsin D occur prior to activation of a mitochondrial pathway of macrophage apoptosis. Pharmacological inhibition or knockout of cathepsin D during pneumococcal infection blocked macrophage apoptosis. As a result of cathepsin D activation, Mcl-1 interacted with its ubiquitin ligase Mule and expression declined. Inhibition of cathepsin D had no effect on early bacterial killing but inhibited the late phase of apoptosis-associated killing of pneumococci in vitro. Mice bearing a cathepsin D-/- hematopoietic system demonstrated reduced macrophage apoptosis in vivo, with decreased clearance of pneumococci and enhanced recruitment of neutrophils to control pulmonary infection. These findings establish an unexpected role for a cathepsin D-mediated lysosomal pathway of apoptosis in pulmonary host defense and underscore the importance of apoptosis-associated microbial killing to macrophage function

Constitutively Active Galpha q and Galpha 13 Trigger Apoptosis through Different Pathways

We investigated the effect of expression of constitutively active Galpha mutants on cell survival. Transfection of constitutively active Galphaq and Galpha13 in two different cell lines caused condensation of genomic DNA and nuclear fragmentation. Endonuclease cleavage of genomic DNA was followed by labeling the DNA fragments and subsequent flow cytometric analysis. The observed cellular phenotype was identical to the phenotype displayed by cells undergoing apoptosis. To distinguish between the apoptosis-inducing ability of the two Galpha-subunits, the signaling pathways involved in this cellular function were investigated. Whereas Galpha q induced apoptosis via a protein kinaseC-dependent pathway, Galpha13 caused programmed cell death through a pathway involving the activation of the small G-protein Rho. Both of the pathways leading to apoptosis were blocked by overexpression of bcl-2. In contrast to other apoptosis-inducing systems, expression of constitutively active Galphaq and Galpha13 triggered apoptosis in high serum as well as in defined medium

The Drosophila Inhibitor of Apoptosis (IAP) DIAP2 Is Dispensable for Cell Survival, Required for the Innate Immune Response to Gram-negative Bacterial Infection, and Can Be Negatively Regulated by the Reaper/Hid/Grim Family of IAP-binding Apoptosis Inducers

Many inhibitor of apoptosis (IAP) family proteins inhibit apoptosis. IAPs contain N-terminal baculovirus IAP repeat domains and a C-terminal RING ubiquitin ligase domain. Drosophila IAP DIAP1 is essential for the survival of many cells, protecting them from apoptosis by inhibiting active caspases. Apoptosis initiates when proteins such as Reaper, Hid, and Grim bind a surface groove in DIAP1 baculovirus IAP repeat domains via an N-terminal IAP-binding motif. This evolutionarily conserved interaction disrupts DIAP1-caspase interactions, unleashing apoptosis-inducing caspase activity. A second Drosophila IAP, DIAP2, also binds Rpr and Hid and inhibits apoptosis in multiple contexts when overexpressed. However, due to a lack of mutants, little is known about the normal functions of DIAP2. We report the generation of diap2 null mutants. These flies are viable and show no defects in developmental or stress-induced apoptosis. Instead, DIAP2 is required for the innate immune response to Gram-negative bacterial infection. DIAP2 promotes cytoplasmic cleavage and nuclear translocation of the NF-{kappa}B homolog Relish, and this requires the DIAP2 RING domain. Increasing the genetic dose of diap2 results in an increased immune response, whereas expression of Rpr or Hid results in down-regulation of DIAP2 protein levels. Together these observations suggest that DIAP2 can regulate immune signaling in a dose-dependent manner, and this can be regulated by IBM-containing proteins. Therefore, diap2 may identify a point of convergence between apoptosis and immune signaling pathways

Bim and Bmf synergize to induce apoptosis in Neisseria gonorrhoeae infection

Abstract: Bcl-2 family proteins including the pro-apoptotic BH3-only proteins are central regulators of apoptotic cell death. Here we show by a focused siRNA miniscreen that the synergistic action of the BH3-only proteins Bim and Bmf is required for apoptosis induced by infection with Neisseria gonorrhoeae (Ngo). While Bim and Bmf were associated with the cytoskeleton of healthy cells, they both were released upon Ngo infection. Loss of Bim and Bmf from the cytoskeleton fraction required the activation of Jun-N-terminal kinase-1 (JNK-1), which in turn depended on Rac-1. Depletion and inhibition of Rac-1, JNK-1, Bim, or Bmf prevented the activation of Bak and Bax and the subsequent activation of caspases. Apoptosis could be reconstituted in Bim-depleted and Bmf-depleted cells by additional silencing of antiapoptotic Mcl-1 and Bcl-XL, respectively. Our data indicate a synergistic role for both cytoskeletal-associated BH3-only proteins, Bim, and Bmf, in an apoptotic pathway leading to the clearance of Ngo-infected cells. Author Summary: A variety of physiological death signals, as well as pathological insults, trigger apoptosis, a genetically programmed form of cell death. Pathogens often induce host cell apoptosis to establish a successful infection. Neisseria gonorrhoeae (Ngo), the etiological agent of the sexually transmitted disease gonorrhoea, is a highly adapted obligate human-specific pathogen and has been shown to induce apoptosis in infected cells. Here we unveil the molecular mechanisms leading to apoptosis of infected cells. We show that Ngo-mediated apoptosis requires a special subset of proapoptotic proteins from the group of BH3-only proteins. BH3-only proteins act as stress sensors to translate toxic environmental signals to the initiation of apoptosis. In a siRNA-based miniscreen, we found Bim and Bmf, BH3-only proteins associated with the cytoskeleton, necessary to induce host cell apoptosis upon infection. Bim and Bmf inactivated different inhibitors of apoptosis and thereby induced cell death in response to infection. Our data unveil a novel pathway of infection-induced apoptosis that enhances our understanding of the mechanism by which BH3-only proteins control apoptotic cell death

Preventing effect of vitamin E on oocytes apoptosis in morphinetreated mice

Several studies have shown that Morphine Sulfate affects on fertility, embryogenesis and consequent pregnancy loss and ultrastructural alterations of oocytes in animal model. This study was done to determine the effect of morphine sulfate on oocytes apoptosis and preventive role of daily supplementation of Vitamin E on oocytes apoptosis in morphine sulfate -treated mice. Twenty-four NMARI female mice were randomly allocated into four experimental groups. For 15 days, control group received saline (0.2 ml/day by subcutaneous injection), group I Vitamin E (60 mg/kg/day orally), group II Morphine Sulfate (10 mg/kg/day by subcutaneous injection) and group III Morphine Sulfate with Vitamin E (60 mg/kg/day orally). Then, animals were superovulated with PSMG (10 Units) and 10 Unites of HCG. The next day the animals were sacrificed, oocytes were flushed from each fallopian tube. The collected oocytes were subjected to determine apoptosis by Tunnel assay with using Fluorescent Microscope. According to our results, the number of retrieved oocytes were 121, 132, 86 and 114 in control, experimental group I, II and III, respectively. Morphine Sulfate treatment increased apoptosis in oocytes to 17.44% whereas oocytes apoptosis was 4.13% in Controls. Supplementation with Vitamin E in Morphine Sulfate -treated mice reduced the oocytes apoptosis to 7.01%. This study showed that Morphine can increase apoptosis in oocytes and Vitamin E treatment significantly reduces oocytes apoptosis in the Morphine Sulfate -treated mice

Pioglitazone inhibits growth of carcinoid cells and promotes TRAIL-induced apoptosis by induction of p21(waf1/cip1)

Background/Aims: We investigated the effect of the peroxisome proliferator-activated receptor-gamma (PPAR-gamma) agonist pioglitazone on growth and TRAIL-induced apoptosis in carcinoid cells. Methods: Carcinoid cells were incubated without and with pioglitazone. Effects on growth were examined by cell count and cell cycle analysis. p21(waf1/cip1) expression was determined by Western blotting. Cytotoxicity assay was performed by FACS analysis. Results: Pioglitazone suppressed the growth and induced apoptosis of carcinoid cells. Additionally, pioglitazone significantly enhanced carcinoid cell death induced by tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL). The enhancement of TRAIL-induced apoptosis was associated with an upregulation of cyclin-dependent kinase inhibitor p21(waf1/cip1) in pioglitazone-treated carcinoid cells. Importantly, overexpression of p21(waf1/cip1) in carcinoid cells by adenoviral gene transfer of p21 sensitized them to TRAIL-induced apoptosis. Conclusions: These results suggest that pioglitazone inhibits cell growth and sensitizes cells to TRAIL-induced apoptosis by induction of p21(waf1/cip1). Therefore, pioglitazone can be an effective therapeutic adjuvant for the treatment of carcinoid tumors. Copyright (C) 2001 S. Karger AG, Basel