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)

The present study was conducted to see the role of NF-kappaB in virulent (Mycobacterium tuberculosis H37Rv) and avirulent (M. tuberculosis H37Ra) mycobacterial infection in THP-1 cells. To inactivate NF-kappaB, pCMV-IkappaBalphaM dn containing THP-1 cell line was generated which showed marked increase in apoptosis with M. tuberculosis H37Rv and M. tuberculosis H37Ra. Infected THP-1-IkappaBalphaM dn cells showed decrease in mitochondrial membrane potential, cytochrome c release, activation of caspase-3 and enhanced TNF-alpha production. Increase in apoptosis of infected THP-1-IkappaBalphaM dn cells resulted in inhibition of intracellular mycobacterial growth. Differential NF-kappaB activation potential was observed with M. tuberculosis H37Rv and M. tuberculosis H37Ra. Both the strains activated NF-kappaB after 4 h in THP-1 cells however after 48 h only M. tuberculosis H37Rv activated NF-kappaB which lead to up-regulation of bcl-2 family anti-apoptotic member, bfl-1/A1. Our results indicated that NF-kappaB activation may be a determinant factor for the success of virulent mycobacteria within macrophages.
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PMID:Differential expression of NF-kappaB in mycobacteria infected THP-1 affects apoptosis. 1720 71

In order to know how caspases contribute to the intracellular fate of Mycobacterium tuberculosis and host cell death in the infected macrophages, we examined the effect of benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethane (z-VAD-fmk), a broad-spectrum caspase inhibitor, on the growth of M. tuberculosis H37Rv in RAW 264 cells. In the cells treated with z-VAD-fmk, activation of caspase-8, caspase-3/7, and caspase-9 was clearly suppressed, and DNA fragmentation of the infected cells was also reduced. Under this experimental condition, it was found that the treatment markedly inhibited bacterial growth inside macrophages. The infected cells appeared to undergo cell death of the necrosis type in the presence of z-VAD-fmk. We further found that z-VAD-fmk treatment resulted in the generation of intracellular reactive oxygen species (ROS) in the infected cells. By addition of a scavenger of ROS, the host cell necrosis was inhibited and the intracellular growth of H37Rv was significantly restored. Among inhibitors specific for each caspase, only the caspase-9-specific inhibitor enhanced the generation of ROS and induced necrosis of the infected cells. Furthermore, we found that severe necrosis was induced by infection with H37Rv but not H37Ra in the presence of z-VAD-fmk. Caspase-9 activation was also detected in H37Rv-infected cells, but H37Ra never induced such caspase-9 activation. These results indicated that caspase-9, which was activated by infection with virulent M. tuberculosis, contributed to the inhibition of necrosis of the infected host cells, presumably through suppression of intracellular ROS generation.
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PMID:Involvement of caspase-9 in the inhibition of necrosis of RAW 264 cells infected with Mycobacterium tuberculosis. 1740 66

Bronchial airway epithelial cells (BAEpC) are among the first cells to encounter M. tuberculosis following airborne infection. However, the response of BAEpC to M. tuberculosis infection has been little studied. This study investigates the response of a human BAEpC cell line (BEAS-2B) to infection with Mycobacterium bovis Bacille Calmette Guerin (BCG). Cultured human BEAS-2B cells were experimentally infected with BCG. Uninfected BEAS-2B cultures were included as controls. Following infection, BEAS-2B cells were evaluated by various methods at various time points up to 3 days. Cell proliferation was evaluated by cellular bioreduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Distribution of cells along the cell cycle was evaluated by FACS analysis of cellular DNA. Apoptotic cells were identified by cell death ELISA and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling method. Eighty-four apoptosis-relevant genes were screened by PCR gene microarray. Translation of Fas, Fas ligand (Fas-L), and Fas-associated death domain (FADD) were evaluated quantitatively by real-time PCR. Expression of Fas and FADD proteins was evaluated by immunofluorescence and Western blot. Activity of caspase-3 and caspase-8 was evaluated by colorimetric assay of their enzymatic activity. BCG infection of BEAS-2B cells inhibits proliferation, induces cell cycle arrest at the G(0)/G(1) phase, causes apoptosis, modulates transcription of multiple apoptosis-relevant genes, promotes translation of Fas, Fas-L, and FADD, upregulates expression of Fas and FADD proteins, and increases activity of caspase-3 and caspase-8. Infection with BCG does not cause any significant change in the secretion of TGF-beta. The roles of Fas and FADD as mediators of BCG-induced apoptosis in BEAS-2B cells were tested by partial blockade of Fas and FADD expression with silencing RNA. Partial blockade of Fas or FADD expression results in a decreased apoptotic response to BCG infection. In conclusion, BCG induces cell cycle arrest and apoptosis in BEAS-2B cells. BCG induced apoptosis of BEAS-2B cells via the Fas death receptor pathway.
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PMID:Induction of cell cycle arrest and apoptosis by BCG infection in cultured human bronchial airway epithelial cells. 1752 97

Mycobacterium avium subsp. paratuberculosis is the etiologic agent of Johne's disease, a chronic granulomatous enteritis in ruminants. ATP has been reported to induce cell death of macrophages and killing of Mycobacterium species in human and murine macrophages. In this study we investigated the short-term effect of ATP on the viability of M. avium subsp. paratuberculosis-infected bovine mononuclear phagocytes and the bacilli within them. Addition of 5 mM ATP to M. avium subsp. paratuberculosis-infected bovine monocytes resulted in 50% cytotoxicity of bovine monocytes at 24 h. Addition of 2'(3')-O-(4-benzoylbenzoyl) ATP triethylammonium salt (Bz-ATP), which is a longer-lived ATP homologue and purinergic receptor agonist, significantly increased the uptake of YO-PRO, which is a marker for membrane pore activation by P2X receptors. Addition of Bz-ATP also stimulated lactate dehydrogenase release and caspase-3 activity in infected bovine monocytes. Neither ATP nor Bz-ATP reduced the survival of M. avium subsp. paratuberculosis in bovine mononuclear phagocytes. Likewise, addition of ATP or Bz-ATP was cytotoxic to murine macrophage cell lines (RAW 264.7 and J774A.1 cells) but did not affect the intracellular survival of M. avium subsp. paratuberculosis, nor were the numbers of viable Mycobacterium avium subsp. avium or Mycobacterium bovis BCG cells altered in bovine mononuclear phagocytes or J774A.1 cells following ATP or Bz-ATP treatment. These data suggest that extracellular ATP does not induce the killing of intracellular M. avium subsp. paratuberculosis in bovine mononuclear phagocytes.
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PMID:Extracellular ATP is cytotoxic to mononuclear phagocytes but does not induce killing of intracellular Mycobacterium avium subsp. paratuberculosis. 1763 11

The development of granulomas is a major histopathological feature of tuberculosis. Very little information is available concerning the physiology and functions of different cell types in the tuberculous granulomas. The aim of this study was to compare the epithelioid cells (ECs) and multinucleated giant cells (MGCs) in the granulomas caused by Mycobacterium tuberculosis complex organisms. Lymph node biopsies from 30 cases of lymphadenitis were studied for expression of the secreted mycobacterial protein MPT64, caspase 3 as a marker of apoptosis, apoptosis-related proteins (Fas Ligand, Fas and Bax) and inflammatory cytokines (interleukin-10, transforming growth factor-beta (TGF-beta), tumour necrosis factor-alpha and interferon-gamma) by immunohistochemistry. MGCs more often contained M. tuberculosis secretory antigen MPT64 (p < 0.001) and expressed more TGF-beta (p = 0.004) than ECs. The total number of apoptotic MGCs was higher than the number of apoptotic ECs (p = 0.04). Interestingly, there was a significant negative correlation between apoptosis and MPT64 expression in MGCs (r = -0.569, p = 0.003), but not in ECs, implying that the heavy antigen load would lead to inhibition of apoptosis in these cells. When compared with ECs, higher percentage of MGCs expressed Fas Ligand and Fas (p < 0.004). The role of MGCs may thus be different from surrounding ECs and these cells by virtue of higher mycobacterial antigen load, more TGF-beta and reduced apoptosis may contribute towards persistence of infection.
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PMID:Differential expression of mycobacterial antigen MPT64, apoptosis and inflammatory markers in multinucleated giant cells and epithelioid cells in granulomas caused by Mycobacterium tuberculosis. 1826 5

DNA vaccination is a potent means for inducing strong cell-mediated immune responses and protective immunity against viral, bacterial and parasite pathogens in rodents. In an attempt to increase cross-presentation through apoptosis, the DNA-encoding caspase-2 prodomain followed by wild-type or catalytically inactive mutated caspase-3 was inserted into a plasmid encoding the 32 kDa mycolyl transferase (Ag85A) from Mycobacterium tuberculosis. Transient transfection showed that the mutated caspase induced slow apoptosis, normal protein expression and NF-kappaB activation while wild-type caspase induced rapid apoptosis, lower protein expression and no NF-kappaB activation. Ag85A specific antibody production was increased by co-expressing the mutated and decreased by co-expressing the wild-type caspase. Vaccination with pro-apoptotic plasmids triggered more Ag85A specific IFN-gamma producing spleen cells, and more efficient IL-2 and IFN-gamma producing memory cells in spleen and lungs after M. tuberculosis challenge. Compared to DNA-encoding secreted Ag85A, vaccination with DNA co-expressing wild-type caspase increased protection after infection with M. tuberculosis, while vaccination with plasmid co-expressing mutated caspase was not protective, possibly due to the stimulation of IL-6, IL-10 and IL-17A production.
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PMID:Immunogenicity and protective efficacy of a tuberculosis DNA vaccine co-expressing pro-apoptotic caspase-3. 1828 Jun 21

Macrophages provide a first line of defense against Mycobacterium tuberculosis. However, in instances where macrophage activation for killing is suboptimal, M. tuberculosis is capable of surviving intracellularly. IL-32 is a recently described cytokine induced by M. tuberculosis in a variety of cell types including human monocytes and macrophages. In this study, we investigated the biological significance of IL-32 in an in vitro model of M. tuberculosis infection in differentiated THP-1 human macrophages in which IL-32 expression was silenced using stable expression of short hairpin RNA (shRNA). Inhibition of endogenous IL-32 production in THP-1 cells that express one of three distinct shRNA-IL-32 constructs significantly decreased M. tuberculosis induction of TNF-alpha by approximately 60%, IL-1beta by 30-60%, and IL-8 by 40-50% and concomitantly increased the number of cell-associated M. tuberculosis bacteria compared with THP-1 cells stably expressing a scrambled shRNA. In THP-1 cells infected with M. tuberculosis and stimulated with rIL-32, a greater level of apoptosis was observed compared with that with M. tuberculosis infection alone. Obversely, there was significant abrogation of apoptosis induced by M. tuberculosis and a concomitant decrease in caspase-3 activation in cells depleted of endogenous IL-32. rIL-32gamma significantly reduced the number of viable intracellular M. tuberculosis bacteria, which was modestly but significantly abrogated with a caspase-3 inhibitor. We conclude that IL-32 plays a host defense role against M. tuberculosis in differentiated THP-1 human macrophages.
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PMID:IL-32 is a host protective cytokine against Mycobacterium tuberculosis in differentiated THP-1 human macrophages. 2019 Jan 43

Activation of human macrophages infected with Mycobacterium tuberculosis was investigated following exposure to microparticles (MP) possessing high anti-tubercular efficacy in mice. A small set of innate responses (cytokine profiles, NO production, Annexin-V staining and caspase-8, caspase-9 and caspase-3 activities) of differentiated THP-1 cells or human monocyte-derived macrophages infected 1:10 in vitro were compared. Cytokines of THP-1 macrophages were comparable in trends, but not in magnitude, with five human genotypes studied. MP reversed suppression of tumor necrosis factor induced by infection, and transiently upregulated gamma-interferon. Drug-free MP surprisingly induced gamma-interferon, but not tumor necrosis factor. Primary cells responded to MP, regardless of drug content, by upregulation of NO; but THP-1 cells did not respond to drug-free MP. About 19% of infected cells exposed to MP underwent apoptosis compared to approximately 11% cells treated otherwise. Cell death induced by drug-free MP was caspase independent. Intracellular bacterial survival varied between individuals. Untreated infection resulted in survival of 900+/-141 cfu; exposure to soluble drugs, drug-containing and blank microparticles respectively, reduced CFU counts to <10, <10 and 102+/-139. These observations indicate that despite variations in magnitude between cells from different sources, innate responses conducive to killing intracellular bacteria were evoked by inhalable MP.
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PMID:Microparticles induce variable levels of activation in macrophages infected with Mycobacterium tuberculosis. 2038 85

The capacity of infected cells to undergo apoptosis upon insult with a pathogen is an ancient innate immune defense mechanism. Consequently, the ability of persisting, intracellular pathogens such as the human pathogen Mycobacterium tuberculosis (Mtb) to inhibit infection-induced apoptosis of macrophages is important for virulence. The nuoG gene of Mtb, which encodes the NuoG subunit of the type I NADH dehydrogenase, NDH-1, is important in Mtb-mediated inhibition of host macrophage apoptosis, but the molecular mechanism of this host pathogen interaction remains elusive. Here we show that the apoptogenic phenotype of MtbDeltanuoG was significantly reduced in human macrophages treated with caspase-3 and -8 inhibitors, TNF-alpha-neutralizing antibodies, and also after infection of murine TNF(-/-) macrophages. Interestingly, incubation of macrophages with inhibitors of reactive oxygen species (ROS) reduced not only the apoptosis induced by the nuoG mutant, but also its capacity to increase macrophage TNF-alpha secretion. The MtbDeltanuoG phagosomes showed increased ROS levels compared to Mtb phagosomes in primary murine and human alveolar macrophages. The increase in MtbDeltanuoG induced ROS and apoptosis was abolished in NOX-2 deficient (gp91(-/-)) macrophages. These results suggest that Mtb, via a NuoG-dependent mechanism, can neutralize NOX2-derived ROS in order to inhibit TNF-alpha-mediated host cell apoptosis. Consistently, an Mtb mutant deficient in secreted catalase induced increases in phagosomal ROS and host cell apoptosis, both of which were dependent upon macrophage NOX-2 activity. In conclusion, these results serendipitously reveal a novel connection between NOX2 activity, phagosomal ROS, and TNF-alpha signaling during infection-induced apoptosis in macrophages. Furthermore, our study reveals a novel function of NOX2 activity in innate immunity beyond the initial respiratory burst, which is the sensing of persistent intracellular pathogens and subsequent induction of host cell apoptosis as a second line of defense.
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PMID:The type I NADH dehydrogenase of Mycobacterium tuberculosis counters phagosomal NOX2 activity to inhibit TNF-alpha-mediated host cell apoptosis. 2042 51

Two different forms of death are commonly observed when Mycobacterium tuberculosis (Mtb)-infected macrophages die: (i) necrosis, a death modality defined by cell lysis and (ii) apoptosis, a form of death that maintains an intact plasma membrane. Necrosis is a mechanism used by bacteria to exit the macrophage, evade host defenses, and spread. In contrast, apoptosis of infected macrophages is associated with diminished pathogen viability. Apoptosis occurs when tumor necrosis factor activates the extrinsic death domain pathway, leading to caspase-8 activation. In addition, mitochondrial outer membrane permeabilization leading to activation of the intrinsic apoptotic pathway is required. Both pathways lead to caspase-3 activation, which results in apoptosis. We have recently demonstrated that during mycobacterial infection, cell death is regulated by the eicosanoids, prostaglandin E(2) (proapoptotic) and lipoxin (LX)A(4) (pronecrotic). Although PGE(2) protects against necrosis, virulent Mtb induces LXA(4) and inhibits PGE(2) production. Under such conditions, mitochondrial inner membrane damage leads to macrophage necrosis. Thus, virulent Mtb subverts eicosanoid regulation of cell death to foil innate defense mechanisms of the macrophage.
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PMID:Apoptosis is an innate defense function of macrophages against Mycobacterium tuberculosis. 2141 28


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