Apoptosis-inducing factor mediates microglial and neuronal apoptosis caused by pneumococcus.
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Streptococcus pneumoniae is the major cause of bacterial meningitis and it damages the hippocampus by inducing neuronal apoptosis. The blocking of caspases provides only partial protection in experimental meningitis, which suggests that there is an additional apoptotic pathway. A trigger of this pathway is the bacterium itself, as exposure of microglia or neurons to live pneumococci induces rapid apoptosis. In this study, apoptosis was not associated with the activation of caspases-1-10 and was not inhibited by z-VAD-fmk, a broad-spectrum caspase inhibitor. Rather, apoptosis was attributed to damage to mitochondria, which was followed by the release of apoptosis-inducing factor (AIF) from the mitochondria, large-scale DNA fragmentation, and hypodiploidy. Furthermore, intracytoplasmatic microinjection of AIF-specific antiserum markedly impaired pneumococcus-induced apoptosis. These findings indicate that AIF may play a central role in brain cell apoptosis and bacterial pathogenesis. (+info)
Productive HIV-1 infection of primary CD4+ T cells induces mitochondrial membrane permeabilization leading to a caspase-independent cell death.
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We have explored in vitro the mechanism by which human immunodeficiency virus, type 1 (HIV-1) induces cell death of primary CD4+ T cells in conditions of productive infection. Although HIV-1 infection primed phytohemagglutinin-activated CD4+ T cells for death induced by anti-CD95 antibody, T cell death was not prevented by a CD95-Fc decoy receptor, nor by decoy receptors of other members of the TNFR family (TNFR1/R2, TRAILR1/R2/OPG, TRAMP) or by various blocking antibodies, suggesting that triggering of death receptors by their cognate ligands is not involved in HIV-induced CD4 T cell death. HIV-1 induced CD4 T cell shrinkage, cell surface exposure of phosphatidylserine, loss of mitochondrial membrane potential (Deltapsim), and mitochondrial release of cytochrome c and apoptosis-inducing factor. A typical apoptotic phenotype (nuclear chromatin condensation and fragmentation) only occurred in around half of the dying cells. Treatment with benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone, a broad spectrum caspase inhibitor, prevented nuclear chromatin condensation and fragmentation in HIV-infected CD4+ T cells and in a cell-free system (in which nuclei were incubated with cytoplasmic extracts from the HIV-infected CD4+ T cells). Nevertheless, benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone did not prevent mitochondrial membrane potential loss and cell death, suggesting that caspases are dispensable for HIV-mediated cell death. Our findings suggest a major role of the mitochondria in the process of CD4 T cell death induced by HIV, in which targeting of Bax to the mitochondria may be involved. (+info)
beta(2)-microglobulin induces apoptosis in HL-60 human leukemia cell line and its multidrug resistant variants overexpressing MRP1 but lacking Bax or overexpressing P-glycoprotein.
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In this study, we examined whether exogenous beta(2)-microglobulin (beta(2)m) can induce apoptosis in the drug sensitive HL-60 leukemia cell line and its drug resistant variants and investigated the molecular mechanism of beta(2)m-induced apoptosis. Our data revealed that beta(2)m is very significantly down-regulated in two multidrug resistant variants of the HL-60 cells: (a) the MRP1-bearing, Bax-deficient HL-60/ADR cell line, and (b) the P-glycoprotein (P-gp) overexpressing HL-60/VCR cell line. However, exogenous beta(2)m induced similar levels of apoptosis in HL-60 cells and these drug resistant variants. beta(2)m-induced apoptosis in HL-60 and HL-60/VCR cells was associated with decreased mitochondrial membrane potential (Deltapsim) but did not affect Deltapsim in HL-60/ADR cells. Surprisingly, cyclosporin A (CsA), a known inhibitor of the mitochondrial permeability transition (MPT) pore, inhibited beta(2)m-induced apoptosis in HL-60/ADR cells but not in HL-60 and HL-60/VCR cells, suggesting that the pro-apoptotic effect of beta(2)m in these cells is not through MPT pore formation. Furthermore, beta(2)m induced the release of cytochrome c and the apoptosis-inducing factor (AIF) from mitochondria in HL-60 and HL-60/VCR cells, but not in HL-60/ADR cells. Additionally, Z-VAD-fmk, a general inhibitor of caspases which inhibited cytochrome c release in HL-60 and HL-60/VCR cells, had no effect on AIF release in any of these cell lines, but inhibited beta(2)m-induced apoptosis in all three cell lines. However, Western blot analysis revealed that caspases-1, -3, -6, -8, and -9 are not activated during beta(2)m-induced apoptosis in these cells. Therefore, beta(2)m-induces apoptosis through an unknown caspase-dependent mitochondrial pathway in HL-60 and HL-60/VCR cells and by a Bax-independent, non-mitochondrial, caspase-dependent pathway in HL-60/ADR cells. (+info)
Pneumococcal pneumolysin and H(2)O(2) mediate brain cell apoptosis during meningitis.
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Pneumococcus is the most common and aggressive cause of bacterial meningitis and induces a novel apoptosis-inducing factor-dependent (AIF-dependent) form of brain cell apoptosis. Loss of production of two pneumococcal toxins, pneumolysin and H(2)O(2), eliminated mitochondrial damage and apoptosis. Purified pneumolysin or H(2)O(2) induced microglial and neuronal apoptosis in vitro. Both toxins induced increases of intracellular Ca(2+) and triggered the release of AIF from mitochondria. Chelating Ca(2+) effectively blocked AIF release and cell death. In experimental pneumococcal meningitis, pneumolysin colocalized with apoptotic neurons of the hippocampus, and infection with pneumococci unable to produce pneumolysin and H(2)O(2) significantly reduced damage. Two bacterial toxins, pneumolysin and, to a lesser extent, H(2)O(2), induce apoptosis by translocation of AIF, suggesting new neuroprotective strategies for pneumococcal meningitis. (+info)
Mitochondrial dysfunction is an essential step for killing of non-small cell lung carcinomas resistant to conventional treatment.
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Apoptosis, a tightly controlled multi-step mechanism of cell death, is important for anti-cancer therapy-based elimination of tumor cells. However, this process is not always efficient. Small cell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC) cells display different susceptibility to undergo apoptosis induced by anticancer treatment. In contrast to SCLC, NSCLC cells are cross-resistant to a broad spectrum of apoptotic stimuli, including receptor stimulation, cytotoxic drugs and gamma-radiation. Since resistance of tumor cells to treatment often accounts for the failure of traditional forms of cancer therapy, in the present study attempts to find a potent broad-range apoptosis inductor, which can kill therapy-resistant NSCLC cells were undertaken and the mechanism of apoptosis induction by this drug was investigated in detail. We found that staurosporine (STS) had cell killing effect on both types of lung carcinomas. Release of cytochrome c, activation of apical and effector caspases followed by cleavage of their nuclear substrates and morphological changes specific for apoptosis were observed in STS-treated cells. In contrast to treatment with radiation or chemotherapy drugs, STS induces mitochondrial dysfunction followed by translocation of AIF into the nuclei. These events preceded the activation of nuclear apoptosis. Thus, in lung carcinomas two cell death pathways, caspase-dependent and caspase-independent, coexist. In NSCLC cells, where the caspase-dependent pathway is less efficient, the triggering of an AIF-mediated caspase-independent mechanism circumvents the resistance of these cells to treatment. (+info)
Apoptotic pathway activation from mitochondria and death receptors without caspase-3 cleavage in failing human myocardium: fragile balance of myocyte survival?
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OBJECTIVES: Activation of the caspase cascade through the mitochondrial and/or death receptor pathway was investigated in the failing human myocardium, in which the mode and extent of the cascade activation are unknown. BACKGROUND: In terminal heart failure, a loss of cardiomyocytes by overload-induced apoptosis is an attractive mechanism, explaining the progressive character of the disease. However, its relevance is unclear, because the specificity of probes for apoptotic deoxyribonucleic acid damage is under debate. METHODS: Left ventricular specimens from 36 explanted failing and 21 nonfailing donor hearts were used for messenger ribonucleic acid detection by semiquantitative reverse-transcription polymerase chain reaction. From these groups, immunoblot analysis was performed in samples from nine failing and six nonfailing donor hearts. RESULTS: In terminally failing hearts, there was a significant accumulation of cytochrome c in the cytosol, which was associated with activation of caspase-9 and downregulation of its inhibitor, caspase-9S. Similarly, the death receptor-induced pathway revealed activation of caspase-8, combined with downregulation of its inhibitors, flice-like inhibitory protein-L (FLIP(L)) and FLIP(S). The unspecific caspase inhibitors, XIAP, hIAP-1 and hIAP-2, were also downregulated. However, the terminal effector caspase-3 was not activated, and its substrate gelsolin, acting in its uncleaved form as a feedback inhibitor of caspase-3, was not cleaved. CONCLUSIONS: In the terminally failing human myocardium, the caspase cascade is partially activated in the presence of a consistent phenotype shift toward enhanced susceptibility to apoptosis. Although the system is still under a fragile control, the partial initiation of the apoptotic program may be of functional relevance also for the surviving cardiomyocytes. (+info)
Roles of caspases in the programmed cell death of motoneurons in vivo.
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Cysteine proteases comprising the caspase family have been considered one of the major executioners of programmed cell death. However, detailed analyses of the programmed cell death of developing motoneurons in mice following the genetic deletion of two key caspases, casp-3 and casp-9, and in the chick embryo following treatment with caspase inhibitors, indicate that normal amounts of cell loss occur although the death process is delayed. Motoneurons undergoing programmed cell death without caspase activities exhibit a nonapoptotic morphology in which nuclear changes such as chromatin condensation are absent or reduced and which exhibit extensive cytoplasmic vacuolization such as is rarely observed in degenerating control neurons. These results suggest that caspases are involved in, but are not indispensable for, the developmental death of motoneurons, and that one function of caspases may be to facilitate the removal of cells that are destined to die. Possible alternative caspase-independent pathways for the programmed death of motoneurons are discussed. (+info)
Wild-type p53 induced sensitization of mutant p53 TNF-resistant cells: role of caspase-8 and mitochondria.
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In the present study, we have investigated the mechanisms by which the restoration of wild-type (wt) p53 functions in p53 mutant cells increases their susceptibility to the cytotoxic action of tumor necrosis factor (TNF). Our data indicate that the resistance of p53-mutated cl.1001 cells to TNF-induced cell death was not due to a defect in the expression of TRADD and FADD, yet correlated with a reduced caspase-8 activation as well as a deficient mitochondrial membrane permeabilization. Moreover, cl.1001 cells failed to translocate the mitochondrial AIF and cytochrome c to the nucleus and to the cytosol, respectively, in response to TNF. Sensitization of these cells, following infection with a recombinant adenovirus encoding wtp53, to TNF-induced cytotoxicity resulted in the restoration of caspase-8 cleavage and the reestablishment of mitochondrial signs of apoptosis. These findings suggest that the cross-talk between p53 and TNF-induced cell death depends on mitochondria and that the combination of TNF and Adwtp53 may be a potential strategy to sensitize mutant p53 TNF-resistant tumors to the cytotoxic action of this cytokine. (+info)