Abstract 371: Mitochondrial Antiviral Signaling (MAVS) Protects Cardiomyocytes Under Oxidative Stress by Interfering with Bax-Mediated Cell Death

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Toshitaka Yajima ◽  
Stanley Park ◽  
Hanbing Zhou ◽  
Michinari Nakamura ◽  
Mitsuyo Machida ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Death ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Cytochrome C Release ◽  
Antiviral Signaling ◽  
Cell Death Pathway ◽  
The Impact

MAVS is a mitochondrial outer membrane protein that activates innate antiviral signaling by recognizing cytosolic viral RNAs and DNAs. While the discovery of MAVS is the first molecular evidence that links mitochondria to innate immune mechanisms, it is still unclear whether MAVS affects mitochondrial cell death as a member of caspase activation and recruitment domain (CARD)-containing proteins. We found that MAVS interacts with Bax through CARD by Yeast two-hybrid and a series of immunoprecipitation (IP) assay, which led us to hypothesize that MAVS functions not only in the innate antiviral mechanisms but also in the mitochondrial cell death pathway. Methods: 1) We examined molecular interaction between MAVS and Bax under oxidative stress by IP using isolated myocytes with H2O2 stimulation and the heart post ischemia-reperfusion (I/R). 2) We evaluated the effect of MAVS on mitochondrial membrane potential and apoptosis under H2O2 stimulation using isolated myocytes with adenoviral MAVS knockdown. 3) We investigated the impact of MAVS on %myocardial infarction (%MI) post I/R using cardiac-specific MAVS knockout (cKO) and transgenic (cTg) mice which we have originally generated. 4) We examined the effect of MAVS on recombinant Bax (rBax)-mediated cytochrome c release using isolated mitochondria from wild type (WT) and MAVS KO mice. Results: 1) The amount of Bax pulled down with MAVS was significantly increased in isolated myocytes with 0.2 mM H2O2 compared to those without stimulation (mean±SD; 1.808±0.14, n=5, p<0.001) and in the heart post I/R compared to sham (2.2±1.19, n=3, p=0.0081). 2) Myocytes with MAVS knockdown showed clear abnormalities in mitochondrial membrane potential and caspace-3 cleavage with 0.2 mM H2O2 compared to control cardiomyocytes. 3) MAVS cKO had significantly larger %MI than WT (81.9 ± 5.8% vs. 42.6 ± 13.6%, n=8, p=0.0008). In contrast, MAVS cTg had significantly smaller %MI that WT (30.0 ± 4.8% vs. 49.2 ± 4.8%, n=10, p=0.0113). 4) Mitochondria from MAVS KO exhibited cytochrome c release after incubation with 2.5 μ g of rBax while those from WT required 10 μ g of rBax. Conclusion: These results demonstrate that MAVS protects cardiomyocyte under oxidative stress by interfering with Bax-mediated cytochrome c release from mitochondria.

Contrasting Role of Concentration in Rivaroxaban Induced Toxicity and Oxidative Stress in Isolated Kidney Mitochondria

Drug Research ◽  
2019 ◽  
Vol 69 (10) ◽  
pp. 523-527
Author(s):  
Fatemeh Samiei ◽  
Hanieh Sajjadi ◽  
Akram Jamshidzadeh ◽  
Enayatollah Seydi ◽  
Jalal Pourahmad
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Cytochrome C Release ◽  
Rat Kidneys

AbstractRivaroxaban as a small molecule is able to directly and reversibly inhibit the factor Xa. This study was designed to figure out the evaluation effect of rivaroxaban on mitochondria obtained from rat kidneys. We isolated mitochondria from rat kidneys using gradient centrifugation. Then, the toxicity parameters including succinate dehydrogenase (SDH) activity, reactive oxygen species (ROS) formation, mitochondrial swelling, mitochondrial membrane potential (MMP) collapse and cytochrome c release were measured in kidneys mitochondria following the exposure to rivaroxaban. The results showed that rivaroxaban (1.4 and 2.8 mM) raised the reactive oxygen species (ROS) generation, swelling in the mitochondria, collapse in the mitochondrial membrane potential (MMP) and cytochrome c release in the mitochondria isolated from kidneys. While, rivaroxaban at a higher concentration of 5.6 mM showed the opposite effect compared to other lower concentrations. The results indicate that rivaroxaban may have antioxidant effects at high concentrations. The results suggest that rivaroxaban (5.6 mM) has protective effects against oxidative stress and mitochondrial toxicity.

scholarly journals Protein Kinase Cδ Targets Mitochondria, Alters Mitochondrial Membrane Potential, and Induces Apoptosis in Normal and Neoplastic Keratinocytes When Overexpressed by an Adenoviral Vector

1999 ◽  
Vol 19 (12) ◽  
pp. 8547-8558 ◽  
Author(s):  
Luowei Li ◽  
Patricia S. Lorenzo ◽  
Krisztina Bogi ◽  
Peter M. Blumberg ◽  
Stuart H. Yuspa
Keyword(s):  
Cell Death ◽  
Membrane Potential ◽  
Protein Kinase ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Fluorescent Protein ◽  
Protein Fusion ◽  
Cell Death Pathway ◽  
Morphological Criteria ◽  
Protein Kinase Cδ

ABSTRACT Inactivation of protein kinase Cδ (PKCδ) is associated with resistance to terminal cell death in epidermal tumor cells, suggesting that activation of PKCδ in normal epidermis may be a component of a cell death pathway. To test this hypothesis, we constructed an adenovirus vector carrying an epitope-tagged PKCδ under a cytomegalovirus promoter to overexpress PKCδ in normal and neoplastic keratinocytes. While PKCδ overexpression was detected by immunoblotting in keratinocytes, the expression level of other PKC isozymes, including PKCα, PKCɛ, PKCζ, and PKCη, did not change. Calcium-independent PKC-specific kinase activity increased after infection of keratinocytes with the PKCδ adenovirus. Activation of PKCδ by 12-O-tetradecanoylphorbol-13-acetate (TPA) at a nanomolar concentration was lethal to normal and neoplastic mouse and human keratinocytes overexpressing PKCδ. Lethality was inhibited by PKC selective inhibitors, GF109203X and Ro-32-0432. TPA-induced cell death was apoptotic as evidenced by morphological criteria, TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) assay, DNA fragmentation, and increased caspase activity. Subcellular fractionation indicated that PKCδ translocated to a mitochondrial enriched fraction after TPA activation, and this finding was confirmed by confocal microscopy of cells expressing a transfected PKCδ-green fluorescent protein fusion protein. Furthermore, activation of PKCδ in keratinocytes altered mitochondrial membrane potential, as indicated by rhodamine-123 fluorescence. Mitochondrial inhibitors, rotenone and antimycin A, reduced TPA-induced cell death in PKCδ-overexpressing keratinocytes. These results indicate that PKCδ can initiate a death pathway in keratinocytes that involves direct interaction with mitochondria and alterations of mitochondrial function.

scholarly journals Caspase Inhibition Extends the Commitment to Neuronal Death Beyond Cytochrome c Release to the Point of Mitochondrial Depolarization

2000 ◽  
Vol 150 (1) ◽  
pp. 131-144 ◽  
Author(s):  
Mohanish Deshmukh ◽  
Keisuke Kuida ◽  
Eugene M. Johnson
Keyword(s):  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Membrane Potential ◽  
Neuronal Death ◽  
Mitochondrial Membrane ◽  
Sympathetic Neurons ◽  
Caspase Inhibitor ◽  
Caspase 9 ◽  
Cytochrome C Release ◽  
Caspase Inhibition

Nerve growth factor (NGF) deprivation induces a Bax-dependent, caspase-dependent programmed cell death in sympathetic neurons. We examined whether the release of cytochrome c was accompanied by the loss of mitochondrial membrane potential during sympathetic neuronal death. NGF- deprived, caspase inhibitor–treated mouse sympathetic neurons maintained mitochondrial membrane poten-tial for 25–30 h after releasing cytochrome c. NGF- deprived sympathetic neurons became committed to die, as measured by the inability of cells to be rescued by NGF readdition, at the time of cytochrome c release. In the presence of caspase inhibitor, however, this commitment to death was extended beyond the point of cytochrome c release, but only up to the subsequent point of mitochondrial membrane potential loss. Caspase-9 deficiency also arrested NGF-deprived sympathetic neurons after release of cytochrome c, and permitted these neurons to be rescued with NGF readdition. Commitment to death in the NGF-deprived, caspase- 9–deficient sympathetic neurons was also coincident with the loss of mitochondrial membrane potential. Thus, caspase inhibition extended commitment to death in trophic factor–deprived sympathetic neurons and allowed recovery of neurons arrested after the loss of cytochrome c, but not beyond the subsequent loss of mitochondrial membrane potential.

scholarly journals Apoptosis Effect of Girinimbine Isolated fromMurraya koenigiion Lung Cancer CellsIn Vitro

2013 ◽  
Vol 2013 ◽  
pp. 1-12 ◽  
Author(s):  
Syam Mohan ◽  
Siddig Ibrahim Abdelwahab ◽  
Shiau-Chuen Cheah ◽  
Mohd Aspollah Sukari ◽  
Suvitha Syam ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cell Death ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Annexin V ◽  
Murraya Koenigii ◽  
Plasma Membrane Permeability ◽  
Lysosomal Stability

Murraya koenigiiSpreng has been traditionally claimed as a remedy for cancer. The current study investigated the anticancer effects of girinimbine, a carbazole alkaloid isolated fromMurraya koenigiiSpreng, on A549 lung cancer cells in relation to apoptotic mechanistic pathway. Girinimbine was isolated fromMurraya koenigiiSpreng. The antiproliferative activity was assayed using MTT and the apoptosis detection was done by annexin V and lysosomal stability assays. Multiparameter cytotoxicity assays were performed to investigate the change in mitochondrial membrane potential and cytochrome c translocation. ROS, caspase, and human apoptosis proteome profiler assays were done to investigate the apoptotic mechanism of cell death. The MTT assay revealed that the girinimbine induces cell death with an IC50of 19.01 μM. A significant induction of early phase of apoptosis was shown by annexin V and lysosomal stability assays. After 24 h treatment with 19.01 μM of girinimbine, decrease in the nuclear area and increase in mitochondrial membrane potential and plasma membrane permeability were readily visible. Moreover the translocation of cytochrome c also was observed. Girinimbine mediates its antiproliferative and apoptotic effects through up- and downregulation of apoptotic and antiapoptotic proteins. There was a significant involvement of both intrinsic and extrinsic pathways. Moreover, the upregulation of p53 as well as the cell proliferation repressor proteins, p27 and p21, and the significant role of insulin/IGF-1 signaling were also identified. Moreover the caspases 3 and 8 were found to be significantly activated. Our results taken together indicated that girinimbine may be a potential agent for anticancer drug development.

scholarly journals Ligands of the Mitochondrial 18 kDa Translocator Protein Attenuate Apoptosis of Human Glioblastoma Cells Exposed to Erucylphosphohomocholine

2008 ◽  
Vol 30 (5) ◽  
pp. 435-450
Author(s):  
Wilfried Kugler ◽  
Leo Veenman ◽  
Yulia Shandalov ◽  
Svetlana Leschiner ◽  
Ilana Spanier ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Cytochrome C ◽  
Cyclosporin A ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Translocator Protein ◽  
Glioblastoma Cells ◽  
Caspase 9 ◽  
Cytochrome C Release ◽  
Induced Apoptosis

Background: We have previously shown that the anti-neoplastic agent erucylphosphohomocholine (ErPC3) requires the mitochondrial 18 kDa Translocator protein (TSPO), formerly known as the peripheral-type benzodiazepine receptor (PBR), to induce cell death via the mitochondrial apoptosis pathway.Methods: With the aid of the dye JC-1 and cyclosporin A, applied to glioblastoma cells, we now investigated the significance of opening of the mitochondrial permeability transition pore (MPTP) for ErPC3-induced apoptosis in interaction with the TSPO ligands, PK 11195 and Ro5 4864. Furthermore, we measured cytochrome c release, and caspase-9 and -3 activation in this paradigm.Results: The human glioblastoma cell lines, U87MG, A172 and U118MG express the MPTP-associated TSPO, voltage-dependent anion channel and adenine nucleotide transporter. Indeed, ErPC3-induced apoptosis was inhibited by the MPTP blocker cyclosporin A and by PK 11195 and Ro5 4864 in a concentration-dependent manner. Furthermore, PK 11195 and Ro5 4864 inhibited collapse of the mitochondrial membrane potential, cytochrome c release, and caspase-9 and -3 activation caused by ErPC3 treatment.Conclusions: This study shows that PK 11195 and Ro5 4864 inhibit the pro-apoptotic function of ErPC3 by blocking its capacity to cause a collapse of the mitochondrial membrane potential. Thus, the TSPO may serve to open the MPTP in response to anti-cancer drugs such as ErPC3.

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