Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P42574 (caspase-3)
 45,978 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In human and rodent macrophages, activation of the P2X7 nucleotide receptor stimulates interleukin-1beta processing and release, apoptosis, and killing of intracellular Mycobacterium tuberculosis. Signaling pathways downstream of this ionotropic ATP receptor are poorly understood. Here we describe the rapid activation of the stress-activated protein kinase (SAPK)/JNK pathway in BAC1 murine macrophages stimulated by extracellular ATP. Brief exposure of the cells to ATP (10-30 min) was sufficient to trigger a rapid accumulation of activated SAPK that was then sustained for >120 min. Several observations indicated that the P2X7 receptor mediated this effect. 1) ATP and 3'-O-(4-benzoyl)benzoyl-ATP were the only agonistic nucleotides. 2) The effect was inhibited by oxidized ATP and the isoquinoline KN-62, two known P2X7 receptor antagonists. 3) ATP-induced SAPK activation could be recapitulated in P2X7 receptor-transfected HEK293 cells, but not in wild-type HEK293 cells. Because P2X7 receptor stimulation can rapidly activate caspase family proteases that have been implicated in the induction of the SAPK pathway, we investigated whether ATP-dependent SAPK activation involved such proteases. Brief exposure of BAC1 macrophages to extracellular ATP induced DNA fragmentation, alpha-fodrin breakdown, and elevated levels of caspase-3-type activity. Asp-Glu-Val-Asp-cho, a caspase-3 inhibitor, inhibited ATP-induced DNA fragmentation and alpha-fodrin proteolysis, but had no effect on ATP-induced SAPK activation. Tyr-Val-Ala-Asp-chloromethyl ketone, a caspase-1 inhibitor, prevented ATP-induced release of processed interleukin-1beta, but not ATP-dependent SAPK activity. We conclude that activation of ionotropic P2X7 nucleotide receptors triggers a strong activation of SAPK via a pathway independent of caspase-1- or caspase-3-like proteases.
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PMID:Stress-activated protein kinase/JNK activation and apoptotic induction by the macrophage P2X7 nucleotide receptor. 1085 31

Macrophage (MPhi) apoptosis, an important innate microbial defense mechanism induced by Mycobacterium tuberculosis (Mtb) H37Ra, depends on the induction of TNF-alpha synthesis. When protein synthesis is blocked, both infection with Mtb and addition of TNF-alpha are required to induce caspase 9 activation, caspase 3 activation and apoptosis. In this study, we show that the second protein synthesis-independent signal involves activation of group IV cytosolic phospholipase A2 (cPLA2). Apoptosis of Mtb-infected MPhi and concomitant arachidonic acid release are abrogated by group IV cPLA2 inhibitors (methyl arachidonyl fluorophosphate and methyl trifluoromethyl ketone), but not by inhibitors of group VI Ca2+-independent (iPLA2; bromoenol lactone) or of secretory low molecular mass PLA2. In MPhi homogenates, the predominant PLA2 activity showed the same inhibitor sensitivity pattern and preferred arachidonic acid over palmitic acid in substrates, also indicating the presence of one or more group IV cPLA2 enzymes. In concordance with these findings, MPhi lysates contained transcripts and protein for group IV cPLA2-alpha and cPLA2-gamma. Importantly, group IV cPLA2 inhibitors significantly reduced MPhi antimycobacterial activity and addition of arachidonic acid, the major product of group IV cPLA2, to infected MPhi treated with cPLA2 inhibitors completely restored the antimycobacterial activity. Importantly, addition of arachidonic acid alone to infected MPhi significantly reduced the mycobacterial burden. These findings indicate that Mtb induces MPhi apoptosis by independent signaling through at least two pathways, TNF-alpha and cPLA2, which are both also critical for antimycobacterial defense of the MPhi.
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PMID:Cytosolic phospholipase A2 participates with TNF-alpha in the induction of apoptosis of human macrophages infected with Mycobacterium tuberculosis H37Ra. 1139 May

In addition to direct bactericidal activities, such as phagocytosis and generation of reactive oxygen species (ROS), neutrophils can regulate the inflammatory response by undergoing apoptosis. We found that infection of human neutrophils with Mycobacterium tuberculosis (Mtb) induced rapid cell death displaying the characteristic features of apoptosis such as morphologic changes, phosphatidylserine exposure, and DNA fragmentation. Both a virulent (H37Rv) and an attenuated (H37Ra) strain of Mtb were equally effective in inducing apoptosis. Pretreatment of neutrophils with antioxidants or an inhibitor of NADPH oxidase markedly blocked Mtb-induced apoptosis but did not affect spontaneous apoptosis. Activation of caspase-3 was evident in neutrophils undergoing spontaneous apoptosis, but it was markedly augmented and accelerated during Mtb-induced apoptosis. The Mtb-induced apoptosis was associated with a speedy and transient increase in expression of Bax protein, a proapoptotic member of the Bcl-2 family, and a more prominent reduction in expression of the antiapoptotic protein Bcl-x(L). Pretreatment with an inhibitor of NADPH oxidase distinctly suppressed the Mtb-stimulated activation of caspase-3 and alteration of Bax/Bcl-x(L) expression in neutrophils. These results indicate that infection with Mtb causes ROS-dependent alteration of Bax/Bcl-x(L) expression and activation of caspase-3, and thereby induces apoptosis in human neutrophils. Moreover, we found that phagocytosis of Mtb-induced apoptotic neutrophils markedly increased the production of proinflammatory cytokine TNF-alpha by human macrophages. Therefore, the ROS-dependent apoptosis in Mtb-stimulated neutrophils may represent an important host defense mechanism aimed at selective removal of infected cells at the inflamed site, which in turn aids the functional activities of local macrophages.
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PMID:Mycobacterium tuberculosis promotes apoptosis in human neutrophils by activating caspase-3 and altering expression of Bax/Bcl-xL via an oxygen-dependent pathway. 1205 53

Mycobacterium tuberculosis (MTB) can induce apoptosis in monocytes/macrophages both in vitro and in vivo, and this phenomenon is associated with mycobacterial survival. The present study addresses the possibility that apoptotic and inflammatory pathways could coexist through a caspase-1-mediated mechanism. In this context, a caspase-1 inhibitor (YVAD), but not caspase-3 (DEVD) or caspase-4 (LEVD) inhibitors, was able to strongly inhibit MTB-induced apoptosis. Moreover, caspase-1 activity was confirmed by the increased maturation of interleukin (IL)-1beta. Of interest, IL-1beta and tumor necrosis factor (TNF)-alpha were produced massively in the course of infection, and both were inhibited by YVAD pretreatment. To determine whether TNF-alpha was produced actively by apoptotic cells, the intracytoplasmatic cytokine content and apoptotic phenotype were analyzed at the single-cell level. Results showed a progressive increase of TNF-alpha production in annexin V-positive cells. These results indicate that MTB-induced apoptosis is associated with proinflammatory cytokine production.
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PMID:Proinflammatory cytokines in the course of Mycobacterium tuberculosis-induced apoptosis in monocytes/macrophages. 1240 97

We previously reported that Mycobacterium tuberculosis infection primes human alveolar macrophages (HAM) for tumor necrosis factor alpha (TNF-alpha)-mediated apoptosis and that macrophage apoptosis is associated with killing internalized bacilli. Virulent mycobacterial strains elicit much less apoptosis than attenuated strains, implying that apoptosis is a defense against intracellular infection. The present study evaluated the potential for phorbol myristate acetate-differentiated THP-1 cells to mimic this response of primary macrophages. Consistent with the behavior of alveolar macrophages, attenuated M. tuberculosis H37Ra and Mycobacterium bovis BCG strongly induce THP-1 apoptosis, which requires endogenous TNF. THP-1 apoptosis is associated with reduced viability of infecting BCG. In contrast, virulent wild-type M. tuberculosis H37Rv and M. bovis do not increase THP-1 apoptosis over baseline. BCG induced early activation of caspase 10 and 9, followed by caspase 3. In contrast, wild-type M. bovis infection failed to activate any caspases in THP-1 cells. BCG-induced THP-1 apoptosis is blocked by retroviral transduction with vectors expressing crmA but not bcl-2. We conclude that differentiated THP-1 cells faithfully model the apoptosis response of HAM. Analysis of the THP-1 cell response to infection with virulent mycobacteria suggests that TNF death signals are blocked proximal to initiator caspase activation, at the level of TNF receptor 1 or its associated intracytoplasmic adaptor complex. Interference with TNF death signaling may be a virulence mechanism that allows M. tuberculosis to circumvent innate defenses leading to apoptosis of infected host cells.
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PMID:THP-1 cell apoptosis in response to Mycobacterial infection. 1249 73

Mycobacterium tuberculosis interacts with macrophages and epithelial cells in the alveolar space of the lung, where it is able to invade and replicate in both cell types. M. tuberculosis-associated cytotoxicity to these cells has been well documented, but the mechanisms of host cell death are not well understood. We examined the induction of apoptosis and necrosis of human macrophages (U937) and type II alveolar epithelial cells (A549) by virulent (H37Rv) and attenuated (H37Ra) M. tuberculosis strains. Apoptosis was determined by both enzyme-linked immunosorbent assay (ELISA) and TdT-mediated dUTP nick end labelling (TUNEL) assay, whereas necrosis was evaluated by the release of lactate dehydrogenase (LDH). Both virulent and attenuated M. tuberculosis induced apoptosis in macrophages; however, the attenuated strain resulted in significantly more apoptosis than the virulent strain after 5 days of infection. In contrast, cytotoxicity of alveolar cells was the result of necrosis, but not apoptosis. Although infection with M. tuberculosis strains resulted in apoptosis of 14% of the cells on the monolayer, cell death associated with necrosis was observed in 59% of alveolar epithelial cells after 5 days of infection. Infection with M. tuberculosis suppressed apoptosis of alveolar epithelial cells induced by the kinase inhibitor, staurosporine. Because our findings suggest that M. tuberculosis can modulate the apoptotic response of macrophages and epithelial cells, we carried out an apoptosis pathway-specific cDNA microarray analysis of human macrophages and alveolar epithelial cells. Whereas the inhibitors of apoptosis, bcl-2 and Rb, were upregulated over 2.5-fold in infected (48 h) alveolar epithelial cells, the proapoptotic genes, bad and bax, were downregulated. The opposite was observed when U937 macrophages were infected with M. tuberculosis. Upon infection of alveolar epithelial cells with M. tuberculosis, the generation of apoptosis, as determined by the expression of caspase-1, caspase-3 and caspase-10, was inhibited. Inhibition of replication of intracellular bacteria resulted in an increase in apoptosis in both cell types. Our results showed that the differential induction of apoptosis between macrophages and alveolar epithelial cells represents specific strategies of M. tuberculosis for survival in the host.
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PMID:Mycobacterium tuberculosis infection causes different levels of apoptosis and necrosis in human macrophages and alveolar epithelial cells. 1292 34

Macrophage apoptosis plays a role in mycobacterial infection. To define the mechanism by which virulent Mycobacterium tuberculosis escapes apoptosis and killing in macrophages, J774 macrophages were infected with virulent M. tuberculosis H37Rv and attenuated H37Ra strains. H37Rv induced less apoptosis than H37Ra, and caspase 3 was activated in H37Ra- and H37Rv-infected macrophages. Intracellular H37Rv bacilli were released at a higher rate into the supernatant than were H37Ra by the sixth day of infection, and this was simultaneously accompanied by the increased necrosis of infected cells showing lactate dehydrogenase (LDH) release. Fas mRNA expression was downregulated and FasL was upregulated in H37Ra- and H37Rv-infected macrophages, while Bcl-2 was upregulated in H37Rv-infected macrophages but downregulated in H37Ra-infected macrophages as seen by real-time PCR. These results indicate that M. tuberculosis H37Ra and H37Rv proliferate in macrophages by preventing them from inducing apoptosis during the early phase of infection, and that M. tuberculosis H37Rv-infected macrophages are found to express Bcl-2 mRNA, which leads to anti-apoptotic activity, and that relatively distinct necrosis might occur during the later phase of infection.
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PMID:Survival of virulent Mycobacterium tuberculosis involves preventing apoptosis induced by Bcl-2 upregulation and release resulting from necrosis in J774 macrophages. 1617 39

We previously reported that macrophage exposure to attenuated strains of pathogenic mycobacteria at multiplicities of infection (MOI) < or = 10 triggers TNF-alpha-mediated apoptosis which reduces the viability of intracellular bacilli. Virulent strains were found to suppress macrophage apoptosis, and it was proposed that apoptosis is an innate defense against intracellular Mycobacterium tuberculosis analogous to apoptosis of virus-infected cells. The potential similarity of host cell responses to intracellular infection with mycobacteria and viruses suggests that M. tuberculosis might lyse infected macrophage when that niche is no longer needed. To investigate this question, we challenged murine macrophages with high intracellular bacillary loads. A sharp increase in cytolysis within 24 h was observed at MOI > or = 25. The primary death mode was apoptosis, based on nuclear morphology and phosphatidyl serine exposure, although the apoptotic cells progressed rapidly to necrosis. Apoptosis at high MOI differs markedly from low MOI apoptosis: it is potently induced by virulent M. tuberculosis, it is TNF-alpha-independent, and it does not reduce mycobacterial viability. Caspase inhibitors failed to prevent high MOI apoptosis, and macrophages deficient in caspase-3, MyD88, or TLR4 were equally susceptible as wild type. Apoptosis was reduced in the presence of cathepsin inhibitors, suggesting the involvement of lysosomal proteases in this novel death response. We conclude that the presence of high numbers of intracellular M. tuberculosis bacilli triggers a macrophage cell death pathway that could promote extracellular spread of infection and contribute to the formation of necrotic lesions in tuberculosis.
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PMID:Macrophage apoptosis in response to high intracellular burden of Mycobacterium tuberculosis is mediated by a novel caspase-independent pathway. 1654 64

Mammalian cell entry (Mce1A) protein of Mycobacterium tuberculosis (Mtb) is involved in bacterial entry and survival in macrophages, which has been shown to induce production of tumor necrosis factor alpha (TNF-alpha). It remains unclear whether and how Mce1A functions upon the type I interleukin-1 receptor-associated protein kinase (IRAK-1) and TNF receptor-associated factor 6 (TRAF-6) of important proinflammatory cytokines. In this study, His-tagged Mce1A was expressed and purified. Also, two pieces of small interfering RNA (siRNA) were designed and synthesized by in vitro transcription, which exhibit specific and efficient silencing effect on mce1a expression. Furthermore, RAW 264.7 murine macrophage-like cells were exposed to His-tagged Mce1A or co-transfected with the Mce1A-expressing plasmid and efficient siRNA, and levels of IRAK-1 and TRAF-6 were then determined by Western blot. We show here that Mce1A induces up-regulations of IRAK-1 and TRAF-6 in macrophages in a dose-dependent manner. The level of Caspase-3 closely related with apoptosis was also determined, whereas no changes were observed. These results indicate that Mtb Mce1A protein induces a proinflammatory response in macrophages.
Tuberculosis (Edinb) 2007 May
PMID:Mammalian cell entry protein of Mycobacterium tuberculosis induces the proinflammatory response in RAW 264.7 murine macrophage-like cells. 1704 27

Mycobacterium tuberculosis complex species survive and replicate in phagosomes of the host cell. Cell death (CD) has been highlighted as one of the probable outcomes in this host-pathogen interaction. Previously, our group demonstrated macrophage apoptosis as a consequence of Mycobacterium bovis infection. In this study, we aimed to identify the contribution of apoptotic effector elements in M. bovis-induced CD. Bovine macrophages were either infected with M. bovis (multiplicity of infection, 10:1) or treated with an M. bovis cell extract (CFE). Structural changes compatible with CD were evaluated. Chromatin condensation was increased three times by the CFE. On the other hand, a terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assay demonstrated that levels of DNA fragmentation induced by M. bovis and CFE were 53.7% +/- 24% and 38.9% +/- 14%, respectively, whereas control cells had a basal proportion of 8.9% +/- 4.1%. Rates of DNA fragmentation were unaffected by the presence of the pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp (z-VAD). Cells treated with 100 mug of CFE for 12 h had a fivefold decrease in the level of mitochondrial outer membrane permeabilization compared to that of untreated cells. Neither M. bovis infection nor CFE treatment induced activation of caspase 3, 8, or 9. Translocation of apoptosis-inducing factor (AIF) to the nucleus was identified in 32% +/- 3.5% and 26.3% +/- 4.9% of M. bovis-infected and CFE-treated cells, respectively. Incubation of macrophages with z-VAD prior to infection did not alter the percentage of cells showing AIF translocation. Our data suggest that M. bovis-induced CD in bovine macrophages is caspase independent with AIF participation.
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PMID:Apoptosis-inducing factor participation in bovine macrophage Mycobacterium bovis-induced caspase-independent cell death. 1715 96


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