Mitochondria-dependent apoptosis induced by nanoscale hydroxyapatite in human gastric cancer SGC-7901 cells.
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Nanoscale hydroxyapatite (nano-HAP) has been reported to exhibit anti-cancer effect on several human cancers, but the molecular mechanism of which remains unclear. The aim of this study was to explore the mechanisms by investigating the effects of nano-HAP on human gastric cancer SGC-7901 cells. Our results showed that nano-HAP significantly reduced cell viability, and induced apoptosis in SGC-7901 cells characterized by hypodiploid DNA contents, morphological changes and DNA fragmentation. The increase in apoptosis was accompanied with the increased expression of Bax, a pro-apoptotic protein, and decreased expression of Bcl-2, an anti-apoptotic protein, the decrease of mitochondrial membrane potential and the release of cytochrome c from mitochondria into cytosol. Furthermore, the activation of caspases-3, and -9, but not activation of caspases-8 was induced by nano-HAP. Z-VAD-fmk, a universal caspase inhibitor, dose-dependently inhibited nano-HAP-induced apoptosis. This study demonstrates that nano-HAP inhibits the proliferation of SGC-7901 cells by inducing apoptosis, and the apoptotic pathway of nano-HAP-induced apoptosis is mediated through the mitochondrial-dependent and caspase-dependent pathway. (+info)
Differential expression of NF-kappaB in mycobacteria infected THP-1 affects apoptosis.
(66/2207)
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. (+info)
Biogenesis and dynamics of mitochondria during the cell cycle: significance of 3'UTRs.
(67/2207)
Nowadays, we are facing a renaissance of mitochondria in cancer biology. However, our knowledge of the basic cell biology and on the timing and mechanisms that control the biosynthesis of mitochondrial constituents during progression through the cell cycle of mammalian cells remain largely unknown. Herein, we document the in vivo changes on mitochondrial morphology and dynamics that accompany cellular mitosis, and illustrate the following key points of the biogenesis of mitochondria during progression of liver cells through the cycle: (i) the replication of nuclear and mitochondrial genomes is synchronized during cellular proliferation, (ii) the accretion of OXPHOS proteins is asynchronously regulated during proliferation being the synthesis of beta-F1-ATPase and Hsp60 carried out also at G2/M and, (iii) the biosynthesis of cardiolipin is achieved during the S phase, although full development of the mitochondrial membrane potential (DeltaPsim) is attained at G2/M. Furthermore, we demonstrate using reporter constructs that the mechanism regulating the accretion of beta-F1-ATPase during cellular proliferation is controlled at the level of mRNA translation by the 3'UTR of the transcript. The 3'UTR-driven synthesis of the protein at G2/M is essential for conferring to the daughter cells the original phenotype of the parental cell. Our findings suggest that alterations on this process may promote deregulated beta-F1-ATPase expression in human cancer. (+info)
Vitamin B6 suppresses apoptosis of NM-1 bovine endothelial cells induced by homocysteine and copper.
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Hyperhomocysteinemia is an important risk factor for atherosclerosis. We previously reported that formation of early atherosclerosis in the rat aorta was associated with hyperhomocysteinemia and reduction of antioxidant activity caused by low concentration of vitamin B(6)in vivo. In the present study, we examined effects of vitamin B(6) on apoptosis of bovine endothelial cells (NM-1 cells) treated with homocysteine and copper. Homocysteine and copper induced extracellular hydrogen peroxide, intracellular ROS and cellular lipid peroxide levels. Cell viability was reduced to 30% compared to that of control cells. On the other hand, pyridoxal treatment as well as EDTA treatment increased viability of NM-1 cells treated with homocysteine and copper to about 60%, and significantly decreased extracellular hydrogen peroxide, intracellular ROS and cellular lipid peroxide levels. The treatment of catalase recovered cell viability and reduced the level of extracellular hydrogen peroxide and intracellular ROS. Cell death by homocysteine and copper was confirmed to be due to apoptosis by evaluation of DNA fragmentation and by TUNEL assay. However, apoptosis of NM-1 cells induced by homocysteine and copper was due to a caspase-independent pathway as it was not inhibited by the caspase inhibitor, Z-VAD-fmk. Apoptosis of NM-1 cells induced by homocysteine and copper accompanied with mitochondrial permeability but not cytochrome c release. These results suggest that pyridoxal treatment suppresses apoptosis of NM-1 cells induced by homocysteine and copper, most likely through antioxidant effects. (+info)
Increased longevity and refractoriness to Ca(2+)-dependent neurodegeneration in Surf1 knockout mice.
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Leigh syndrome associated with cytochrome c oxidase (COX) deficiency is a mitochondrial disorder usually caused by mutations of SURF1, a gene encoding a putative COX assembly factor. We present here a Surf1-/- recombinant mouse obtained by inserting a loxP sequence in the open reading frame of the gene. The frequency of -/-, +/+ and +/- genotypes in newborn mice followed a mendelian distribution, indicating that the ablation of Surf1 is compatible with postnatal survival. The biochemical and assembly COX defect was present in Surf1(loxP)-/- mice, but milder than in humans. Surprisingly, not only these animals failed to show spontaneous neurodegeneration at any age, but they also displayed markedly prolonged lifespan, and complete protection from Ca(2+)-dependent neurotoxicity induced by kainic acid. Experiments on primary neuronal cultures showed markedly reduced rise of cytosolic and mitochondrial Ca(2+) in Surf1(loxP)-/- neurons, and reduced mortality, compared to controls. The mitochondrial membrane potential was unchanged in KO versus wild-type neurons, suggesting that the effects of the ablation of Surf1 on Ca(2+) homeostasis, and possibly on longevity, may be independent, at least in part, from those on COX assembly and mitochondrial bioenergetics. (+info)
Mitochondrial c-Jun NH2-terminal kinase prevents the accumulation of reactive oxygen species and reduces necrotic damage in neural tumor cells that lack trophic support.
(70/2207)
In response to different stress signals, the c-Jun NH(2)-terminal kinase (JNK) can trigger cell death. However, JNK also facilitates the survival and cell cycle progression of tumor cells by mechanisms that are poorly defined. Here, we show that schwannoma RN22 cells can survive and proliferate under serum-free conditions although serum withdrawal rapidly induces mitochondrial fission and swelling. Although the morphologic changes observed in the mitochondria did not trigger cytochrome c release, they were accompanied by an increase in the mitochondrial membrane potential (DeltaPsi(M)) and of immunoreactivity for active JNK in these organelles. Pharmacologic inhibition of JNK provoked a further increase of the DeltaPsi(M), an increase in reactive oxygen species (ROS) production, and a sustained decrease in cell viability due to necrosis. This increase in necrosis was prevented by the presence of ROS scavengers. Immunoreactivity for active JNK was also observed in the mitochondria of neuroblastoma 1E-115 and neuroblastoma 2a neuroblastoma cell lines on serum withdrawal, whereas active JNK was barely detected in serum-deprived fibroblasts. Accordingly, the reduction in neural tumor cell viability induced by JNK inhibition was largely attenuated in serum-deprived fibroblasts. These data indicate that local activation of JNK in the mitochondria can protect against necrotic cell death associated with ROS production, facilitating the growth of neural tumor cells subjected to serum deprivation. (+info)
Isoflurane preconditioning uncouples mitochondria and protects against hypoxia-reoxygenation.
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Ischemic cardiac injury can be substantially alleviated by exposing the heart to pharmacological agents such as volatile anesthetics before occurrence of ischemia-reperfusion. A hallmark of this preconditioning phenomenon is its memory, when cardioprotective effects persist even after removal of preconditioning stimulus. Since numerous studies pinpoint mitochondria as crucial players in protective pathways of preconditioning, the aim of this study was to investigate the effects of preconditioning agent isoflurane on the mitochondrial bioenergetic phenotype. Endogenous flavoprotein fluorescence, an indicator of mitochondrial redox state, was elevated to 195 +/- 16% of baseline upon isoflurane application in intact cardiomyocytes, indicating more oxidized state of mitochondria. Isoflurane treatment also elicited partial dissipation of mitochondrial transmembrane potential, which remained depolarized even after anesthetic withdrawal (tetramethylrhodamine fluorescence intensity declined to 83 +/- 3 and 81 +/- 7% of baseline during isoflurane exposure and washout, respectively). Mild uncoupling, with preserved ATP synthesis, was also detected in mitochondria that were isolated from animals that had been previously preconditioned by isoflurane in vivo, revealing its memory nature. These mitochondria, after exposure to hypoxia and reoxygenation, exhibited better preserved respiration and ATP synthesis compared with mitochondria from nonpreconditioned animals. Partial mitochondrial depolarization was paralleled by a diminished Ca(2+) uptake into isoflurane-treated mitochondria, as indicated by the reduced increment in rhod-2 fluorescence when mitochondria were challenged with increased Ca(2+) (180 +/- 24 vs. 258 +/- 14% for the control). In conclusion, isoflurane preconditioning elicits partial mitochondrial uncoupling and reduces mitochondrial Ca(2+) uptake. These effects are likely to reduce the extent of the mitochondrial damage after the hypoxic stress. (+info)
Stimulation-induced changes in NADH fluorescence and mitochondrial membrane potential in lizard motor nerve terminals.
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To investigate mitochondrial responses to repetitive stimulation, we measured changes in NADH fluorescence and mitochondrial membrane potential (Psi(m)) produced by trains of action potentials (50 Hz for 10-50 s) delivered to motor nerve terminals innervating external intercostal muscles. Stimulation produced a rapid decrease in NADH fluorescence and partial depolarization of Psi(m). These changes were blocked when Ca2+ was removed from the bath or when N-type Ca2+ channels were inhibited with omega-conotoxin GVIA, but were not blocked when bath Ca2+ was replaced by Sr2+, or when vesicular release was inhibited with botulinum toxin A. When stimulation stopped, NADH fluorescence and Psi(m) returned to baseline values much faster than mitochondrial [Ca2+]. In contrast to findings in other tissues, there was usually little or no poststimulation overshoot of NADH fluorescence. These findings suggest that the major change in motor terminal mitochondrial function brought about by repetitive stimulation is a rapid acceleration of electron transport chain (ETC) activity due to the Psi(m) depolarization produced by mitochondrial Ca2+ (or Sr2+) influx. After partial inhibition of complex I of the ETC with amytal, stimulation produced greater Psi(m) depolarization and a greater elevation of cytosolic [Ca2+]. These results suggest that the ability to accelerate ETC activity is important for normal mitochondrial sequestration of stimulation-induced Ca2+ loads. (+info)