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At least 721 records · Page 40Linked to original sources
PubMed1998

Mercuric chloride induces apoptosis in human T lymphocytes: evidence of mitochondrial dysfunction.

The major objective of our study was to define the mechanism by which mercuric chloride (HgCl2) induces human T-cell death. Human peripheral blood T-cells were exposed to 0-40 microm HgCl2 and then analyzed for biochemical and molecular features of T-cell apoptosis. HgCl2-treated cells exhibited increased Hoechst 33258 fluorescence while maintaining their ability to exclude the vital stain 7-aminoactinomycin D. To further evaluate cell death and distinguish between apoptosis and necrosis, translocation of phosphatidylserine to the outer layer of the plasma membrane (annexin V binding), DNA fragmentation (TUNEL assay), and cleavage of poly (ADP-ribose) polymerase (PARP) were assessed. In the presence of 20-40 microm HgCl2, T-cells exhibited increased annexin V binding (28%) and DNA fragmentation (31%). HgCl2-dependent PARP cleavage was also observed by Western blot analysis. Because degradative changes associated with apoptosis are often preceded by disruption of mitochondrial function, HgCl2-treated cells were assessed for disruption of the mitochondrial transmembrane potential (DeltaPsim) and development of the mitochondrial permeability transition state. Using DiOC6(3), we demonstrated that HgCl2 exposure resulted in a decrease in the DeltaPsim. Because a decline in DeltaPsim can disturb the intracellular pH (pHi), we used the fluorescent probe, SNARF-1, to assess intracellular acidification. Treatment of T-cells with HgCl2 resulted in reduced pHi from 7.0 to 6.7. Concomitant with these observations, the fluorescent probe, hydroethidine, was utilized to demonstrate that uncoupled mitochondrial electron transport resulted in increased reactive oxygen species (ROS) generation. Interestingly, in spite of these alterations to mitochondrial function, translocation of cytochrome c to the cytosol was not detected; this correlated with enhanced bcl-2 levels in HgCl2-treated cells. In conclusion, HgCl2 exposure results in oxidative stress and activation of death signaling pathways leading to apoptosis. Collectively, our studies indicate that individual mercurial species are capable of inducing T-cell death by activating specific apoptotic cascades.

Apoptosis↗
PubMed2001

Parkinson's disease, pesticides and mitochondrial dysfunction.

Selective nigral degeneration with inclusion formation provoked by systemic administration of the herbicide rotenone, through inhibition of complex I, raises the question of pesticide exposure and environmental factors in general, as a cause of Parkinson's disease (PD). Toxin-induced complex I inhibition probably represents one of many potential causes of PD, but it alerts us to the dangers of such substances in the environment and the need to identify genetically susceptible populations. When vulnerable individuals become known, perhaps they should stay out of the garden.

Animals↗
PubMed2006

Molecular mechanisms of insulin resistance in humans and their potential links with mitochondrial dysfunction.

Recent studies using magnetic resonance spectroscopy have shown that decreased insulin-stimulated muscle glycogen synthesis due to a defect in insulin-stimulated glucose transport activity is a major factor in the pathogenesis of type 2 diabetes. The molecular mechanism underlying defective insulin-stimulated glucose transport activity can be attributed to increases in intramyocellular lipid metabolites such as fatty acyl CoAs and diacylglycerol, which in turn activate a serine/threonine kinase cascade, thus leading to defects in insulin signaling through Ser/Thr phosphorylation of insulin receptor substrate (IRS)-1. A similar mechanism is also observed in hepatic insulin resistance associated with nonalcoholic fatty liver, which is a common feature of type 2 diabetes, where increases in hepatocellular diacylglycerol content activate protein kinase C-epsilon, leading to reduced insulin-stimulated tyrosine phosphorylation of IRS-2. More recently, magnetic resonance spectroscopy studies in healthy lean elderly subjects and healthy lean insulin-resistant offspring of parents with type 2 diabetes have demonstrated that reduced mitochondrial function may predispose these individuals to intramyocellular lipid accumulation and insulin resistance. Further analysis has found that the reduction in mitochondrial function in the insulin-resistant offspring can be mostly attributed to reductions in mitochondrial density. By elucidating the cellular and molecular mechanisms responsible for insulin resistance, these studies provide potential new targets for the treatment and prevention of type 2 diabetes.

Diabetes Mellitus, Type 2↗
PubMed2004

The human TAZ gene complements mitochondrial dysfunction in the yeast taz1Delta mutant. Implications for Barth syndrome.

Barth syndrome is a genetic disorder that is caused by different mutations in the TAZ gene G4.5. The yeast gene TAZ1 is highly homologous to human TAZ, and the taz1Delta mutant has phospholipid defects similar to those observed in Barth syndrome cells, including aberrant cardiolipin species and decreased cardiolipin levels. Subcellular fractionation studies revealed that Taz1p is localized exclusively in mitochondria, which supports the theory that tafazzins are involved in cardiolipin remodeling. Because cardiolipin plays an important role in respiratory function, we measured the energy transformation and osmotic properties of isolated mitochondria from the taz1Delta mutant. Energy coupling in taz1Delta mitochondria was dependent on the rate of oxidative phosphorylation, as coupling was diminished when NADH was used as a respiratory substrate but was unaffected when ethanol was the substrate. Membrane stability was compromised in taz1Delta mitochondria exposed to increased temperature and hypotonic conditions. Mitochondria from taz1Delta also displayed decreased swelling in response to ATP, which induces the yeast mitochondrial unspecific channel, and to alamethicin, a membrane-disrupting agent. Coupling was measured in taz1Delta cells containing different splice variants of the human TAZ gene. Only the variant that restores wild type cardiolipin synthesis (lacking exon 5) restored coupling in hypotonic conditions and at elevated temperature. These findings may shed light on the mitochondrial deficiencies observed in Barth syndrome.

Acyltransferases↗
PubMed1998

Low-level methylmercury exposure causes human T-cells to undergo apoptosis: evidence of mitochondrial dysfunction.

There is growing evidence that heavy metals, in general, and mercurial compounds, in particular, are immunotoxic to the human immune system. The major focus of our study is to demonstrate that methylmercuric chloride (MeHgCl) kills human lymphocytes by inducing apoptosis. T-cells exposed to 0.6-5 microM MeHgCl for 24 h were analyzed by flow cytometry. Methylmercury-treated cells exhibited increased Hoechst 33258 fluorescence while maintaining their ability to exclude the vital stain 7-aminoactinomycin. Furthermore, T-cells exposed to methylmercury exhibited changes in light scatter patterns that included decreased forward light scatter and increased side light scatter. The light scatter and fluorescent changes were consistent with morphological alterations displayed by cells during apoptosis. Cell death was further evaluated by assessing annexin V binding to the plasma membrane. Methylmercury-treated cells exhibited increased annexin V binding indicative of phosphatidylserine translocation to the outer leaflet of the plasma membrane. Using the fluorescent probe DiOC6(3), we noted that methylmercury exposure resulted in a decrease in mitochondrial transmembrane potential (Psim). Since a low Psim is associated with altered mitochondrial function, we also determined if exposure to methylmercury potentiated reactive oxygen species (ROS) generation. We noted that treated cells generated ROS, as evidenced by oxidation of hydroethidine and the generation of the fluorescent product, ethidium. Finally, we evaluated the effect of methylmercury on T-cell GSH content utilizing the fluorescent probe monochlorobimane; in the presence of MeHgCl, there is a marked loss in reduced cell thiols. The results of the study indicate that a key event in the induction of T-cell apoptosis by mercuric compounds is depletion in the thiol reserve which predisposes cells to ROS damage and at the same time activates death signaling pathways.

Apoptosis↗
PubMed1997

Successful treatment by direct hemoperfusion of coma possibly resulting from mitochondrial dysfunction in acute valproate intoxication.

PURPOSE: We evaluated the efficacy of direct hemoperfusion (DHP) for treatment of acute valproate (VPA) intoxication and speculate on the biochemical perturbations that suggest a mechanism of coma induced by VPA overdose. PATIENT AND METHODS: The comatose patient was hospitalized approximately 6 h after ingesting 18 g VPA. DHP, with 200 g activated charcoal, was performed for 6 h. The plasma concentrations of VPA and Glasgow coma scale scores after admission were estimated. Before and after DHP, urine samples were tested in serial fashion for VPA metabolites, organic acids, and acyl carnitine esters of fatty acids. RESULTS: Plasma VPA was efficiently adsorbed on activated charcoal. The patient's plasma concentration of VPA decreased from 471 microg/ml (2,830 microM) to 45 microg/ml (270 microM), at which point the patient became alert. The half-life (t1/2) of VPA was calculated as 4.4 h before DHP and as 1.8 h during DHP. Before DHP, lactate and VPA-glucuronide markedly increased in urine samples, but beta-keto-VPA, a major mitochondrial metabolite, was not detected. Urinary excretion of carnitine esters of medium chain (C8-C10) dicarboxylic acids was increased. After DHP, lactate and VPA-glucuronide decreased, but a significant amount of beta-keto-VPA was demonstrated. Carnitine esters of medium chain dicarboxylic acids were decreased. CONCLUSIONS: DHP with activated charcoal was effective treatment for the patient with acute VPA intoxication and coma. The onset of coma may have been related to inhibition of beta-oxidation in the mitochondria, which was reversible by elimination of plasma VPA by DHP.

Acute Disease↗
PubMed2004

Role of mitochondrial dysfunction in combined bile acid-induced cytotoxicity: the switch between apoptosis and necrosis.

The goal of this investigation was to determine whether chenodeoxycholic acid (CDCA)-induced apoptosis is prevented by ursodeoxycholic acid (UDCA) or tauroursodeoxycholic acid (TUDC) and to characterize the involvement of mitochondria in the process. Cultured human HepG2 cells were treated in a dose- and time-dependent protocol in order to establish a sufficiently low exposure to CDCA that causes apoptosis but not necrosis. Low-dose CDCA induced an S-phase block and G2 arrest of the cell cycle, as determined by flow cytometry. As a result, cell proliferation was inhibited. CDCA-induced apoptosis, as determined by fluorescence microscopy of Hoechst 33342-stained nuclei, was evident upon coincubation with TUDC. Additionally, after exposure to UDCA plus CDCA, the cell membrane was permeable to fluorescent dyes. Caspase-9-like activity, poly(ADP-ribose) polymerase (PARP) cleavage, and extensive DNA fragmentation were detected in CDCA-exposed cells and in cells coincubated with TUDC, but not UDCA. CDCA caused a decrease in mitochondrial membrane potential and depletion of ATP, both of which were potentiated by UDCA but not TUDC. The results suggest that UDCA potentiates CDCA cytotoxicity, probably at the level of induction of the mitochondrial permeability transition (MPT). Consequently, as suggested by the lack of the main hallmarks of the apoptotic pathway, in the presence of UDCA, CDCA-induced apoptosis is not properly executed but degenerates into necrosis.

Adenosine Triphosphate↗
PubMed2002

Amelioration of mitochondrial dysfunction and apoptosis of two-cell mouse embryos after freezing and thawing by the high frequency liquid nitrogen infusion.

Liquid nitrogen (LN2) infusions are currently used in a slow controlled-rate freezing during cryopreservation. The effects of two different LN2 infusion frequencies (conventional, slow 50 infusions/min and high 120 infusions/min) were studied with frozen-thawed two-cell mouse embryos and their subsequent development to blastocysts. The embryos that were subjected to the high frequency LN2 infusion (HFLI) showed a significantly higher survival rate over the low frequency LN2 infusion (LFLI) (50.7 vs. 34.6%, P < 0.05). The blastocyst formation was also higher in HFLI (76.7%) than LFLI (44.0%, P < 0.05) with respective to the number of cells in a blastocyst of 71.6 8.0 (n = 20) and 62.5 +/- 4.7 (n = 20) (P < 0.05). The relative amount of H2O2 in an embryo that was assessed by a fluorescence intensity of 2',7'-dichlorofluorecein (DCF) showed a difference between the procedures with 16.6 +/- 1.6 (n = 21) and 23.4 +/- 1.8 (n = 24) for HFLI and LFLI, respectively (P < 0.05). Mitochondrial staining by Rhodamine 123 showed that the number and distribution of viable mitochondria were similar in both procedures, but fewer mitochondria were observed with a marked aggregation in the arrested embryos, indicating a mitochondrial disintegration. The mitochondrial membrane potential was visualized by a membrane potential-sensitive fluorescent probe, 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1). There was a decrease in the number of mitochondria that had a high membrane potential, and they showed a peripheral redistribution along the cell membrane in LFLI. A fluorescent staining of the actin filaments revealed a discontinuity that was noticeably at the peripheral "actin band" in LFLI. The DNA fragmentation was assessed by the dUTP nick end-labeling (TUNEL). The results showed a higher DNA fragmentation of blastocyst nuclei in LFLI compared to HFLI (65.6 vs. 36.0%, P < 0.05). Based on these observations, it was concluded that HFLI was better than LFLI in the case of freezing the mouse 2-cell embryos for preserving cytoskeletons and mitochondrial integrities. This could subsequently lead to a higher survival and developmental rate of the cryopreserved mouse embryos.

Actins↗
PubMed1987

The effect of dilazep on puromycin-induced rat renal mitochondrial dysfunction.

The effect of tetrahydro-1 H-1,4 (5H)-dipropanol bis(3,4,5-trimethoxybenzoate)hydrochloride monohydrate (dilazep, Comelian) on puromycin-induced rat renal damage was investigated. In vivo study: Rats were divided into 3 groups, the control group; untreated, the puromycin group; puromycin (150 mg/kg) was injected intraperitoneally once, the dilazep + puromycin group; puromycin (150 mg/kg) was injected 1 h after intraperitoneal dilazep injection (2 mg/kg), and dilazep (2 mg/kg) was injected every 12 h until the end of the experiment. In each group, 84 h after puromycin injection, kidneys were isolated and renal mitochondria were prepared. The endogenous phospholipase activity in kidney homogenate was determined by high performance liquid chromatography. The activities of three segments (NADH-cytochrome c reductase, succinate-cytochrome c reductase and cytochrome c oxidase) of the electron-transport chain in mitochondria were measured enzymatically. In the puromycin group, phospholipase activity was increased and activities of all of three segments of the electron-transport chain were decreased. In the dilazep + puromycin group, premedication with dilazep prevented activation of phospholipase and maintained mitochondrial electron-transport activity. In vitro study: Mitochondria prepared from intact rat kidney were incubated with phospholipase C. Activities of the mitochondrial electron-transport chain were deteriorated by phospholipase C. These results indicated that activation of endogenous phospholipase, which digests membrane phospholipids, essential components in maintaining mitochondrial electron-transport activity, is responsible for the puromycin-induced renal damage. Premedication with dilazep prevented the damage by inhibition of the activation of phospholipase.

Animals↗
PubMed1999

Endogenous peroxynitrite mediates mitochondrial dysfunction in rat diaphragm during endotoxemia.

It has been shown that nitric oxide (NO), synthesized by the inducible NO synthase (iNOS) expressed in the diaphragm during endotoxemia, participates in the development of muscular contractile failure. The aim of the present study was to investigate whether this deleterious action of NO was related to its effects on cellular oxidative pathways. Rats were inoculated with E. coli lipopolysaccharide (LPS) or sterile saline solution (controls) and studied at 3 and 6 h after inoculation. iNOS protein and activity could be detected in the rat diaphragm as early as 3 h after LPS, with a sustained steady-state concentration of 0.5 microM NO in the muscle associated with increased detection of hydrogen peroxide (H(2)O(2)). In vitro, the same NO concentration produced a marked increase in H(2)O(2) production by isolated control diaphragm mitochondria, thus reflecting a higher intramitochondrial concentration of nondiffusible superoxide anion (O(2)(-.)). In a similar way, whole diaphragmatic muscle and diaphragm mitochondria from endotoxemic rats showed a progressive increase in H(2)O(2) production associated with uncoupling and decreased phosphorylating capacity. Simultaneous with the maximal impairment in respiration (6 h after LPS), nitration of mitochondrial proteins (a peroxynitrite footprint) was detected and diaphragmatic force was reduced. Functional mitochondrial abnormalities, nitration of mitochondrial proteins, and the decrease in force were significantly attenuated by administration of the NOS inhibitor L-NMMA. These results show that increased and sustained NO levels lead to a consecutive formation of O(2)(-.) that reacts with NO to form peroxynitrite, which in turn impairs mitochondrial function, which probably contributes to the impairment of muscle contractility. during endotoxemia.

Animals↗
PubMed1999

Dysregulation of the cyclin-dependent kinase inhibitor p21WAF1/CIP1/MDA6 increases the susceptibility of human leukemia cells (U937) to 1-beta-D-arabinofuranosylcytosine-mediated mitochondrial dysfunction and apoptosis.

The effects of dysregulation of the cyclin-dependent kinase inhibitor p21WAF1/CIP1 on the apoptotic response of U937 monocytic leukemia cells to 1-beta-D-arabinofuranosylcytosine (ara-C) were examined. After a 6-h exposure to 1 microM ara-C, cells stably transfected with a p21WAF1/CIP1 antisense construct were significantly more sensitive to the induction of classic apoptotic morphology, DNA fragmentation, caspase-3 activation, poly(ADP-ribose) polymerase degradation, and underphosphorylation of the retinoblastoma protein (pRb) than their empty-vector counterparts. Enhanced susceptibility of antisense-expressing cells to ara-C was accompanied by a corresponding reduction in clonogenic and suspension culture growth. The increased sensitivity of these cells to ara-C-mediated lethality could not be attributed to cytokinetic perturbations, nor did ara-CTP formation or (ara-C)DNA incorporation differ significantly between the cell lines. Moreover, synchronization of p21 antisense-expressing cells in S-phase by aphidicolin block resulted in a further increase in ara-C-mediated apoptosis, suggesting enhanced drug sensitivity of the S-phase cell fraction. After exposure to ara-C, p21 antisense-expressing cells displayed a greater decline in mitochondrial membrane potential (deltapsi(m)) and generation of reactive oxygen species than their empty-vector counterparts, as well as early potentiation (e.g., within 2-4 h) of cytochrome c release into the cytosolic S-100 fraction. Lastly, ara-C-mediated increases in mitogen-activated protein kinase activity over basal levels were attenuated in p21 antisense-expressing cells. Collectively, these findings indicate that dysregulation of the cyclin-dependent kinase inhibitor p21WAF1/CIP1 increases the susceptibility of U937 human leukemia cells to ara-C-related lethality, and this phenomenon occurs as a relatively early event that is independent of cell cycle or pharmacodynamic factors and is associated with mitochondrial perturbations implicated in activation of the apoptotic protease cascade.

Antimetabolites, Antineoplastic↗
PubMed1988

Prostaglandin I2 analogue and propranolol prevent ischaemia induced mitochondrial dysfunction through the stabilisation of lysosomal membranes.

Leakage of lysosomal enzymes is associated with irreversible cellular damage. To determine the effect of prostaglandin I2 analogue and propranolol on the ischaemic myocardium in relation to changes in lysosomal integrity 26 anaesthetised mongrel dogs were divided into three treatment groups and subjected to 2 h coronary occlusion. In the control group (n = 12) physiological saline was infused throughout the experiment. In the prostaglandin I2 analogue group (n = 7) the prostaglandin I2 analogue, OP-41483-alpha-CD;5(E)-6-Deoxa-6,9 alpha-methylene-15-cyclopentyl-16,17,18,19,20-pentanor-PGI2. alpha-cyclodextrin clathrate (5 ng.kg-1.min-1) was infused from 25 min before occlusion until the end of the experiment. In the propranolol group (n = 7) propranolol (0.3 mg.kg-1) was injected for 10 min 25 min before occlusion. Two hours after occlusion mitochondria were prepared from both ischaemic and non-ischaemic areas in each group and their function measured polarographically with succinate as substrate. Fractionation of myocardial tissue from both non-ischaemic and ischaemic areas was performed and the activities of lysosomal enzymes (N-acetyl-beta-glucosaminidase; beta-glucuronidase) were measured. In the control group, mitochondrial function in the ischaemic area was reduced compared with that from the non-ischaemic area. The activities of both lysosomal enzymes were increased significantly in the supernatant fraction obtained from the ischaemic area compared with those for the supernatant from the non-ischaemic area. The administration of prostaglandin I2 analogue or propranolol not only prevented the leakage of lysosomal enzymes but also maintained mitochondrial function.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2003

Dimethyl disulfide exerts insecticidal neurotoxicity through mitochondrial dysfunction and activation of insect K(ATP) channels.

The plant-derived insecticides have introduced a new concept in insecticide research. In response to insect attacks, some plants can release volatile sulfur compounds such as dimethyl disulfide (DMDS) in the atmosphere, which are lethal for the generalist insects. We demonstrate that DMDS induced an uncommon complex neurotoxic activity. The studies of in vivo toxicity of DMDS in three insect species and mice indicated a highest bioactivity for insects. Although DMDS did not alter the electrophysiological properties of the cockroach Periplaneta americana giant axon, it affected the synaptic transmission at the presynaptic level resulting in an inhibition of the neurotransmitter release. Whole cell patch-clamp experiments performed on cockroach cultured dorsal unpaired median (DUM) neurons revealed a dose-dependent hyperpolarization induced by DMDS associated with a decrease in the input resistance and the disappearance of action potentials. The hyperpolarization was inhibited by glibenclamide and tolbutamide, and was dependent on intracellular ATP concentration, demonstrating a neurotoxicity via the activation of KATP channels. Finally, the same effects observed with oligomycin, 2,4-dinitrophenol, and KCN together with the studies of DMDS toxicity on isolated mitochondria confirmed an unusual action occurring through an inhibition of the mitochondrial respiratory chain complex IV (cytochrome oxydase). This DMDS-induced inhibition of complex IV subsequently decreased the intracellular ATP concentration, which thereby activated neuronal KATP channels mediating membrane hyperpolarization and reduction of neuronal activity.

Action Potentials↗
PubMed2004

The cellular prion protein (PrPC) prevents apoptotic neuronal cell death and mitochondrial dysfunction induced by serum deprivation.

Prion diseases are transmissible neurodegenerative disorders that are invariably fatal in humans and animals. Although the nature of the infectious agent and pathogenic mechanisms of prion diseases are not clear, it has been reported that prion diseases may be associated with aberrant metabolism of cellular prion protein (PrP(C)). In various reports, it has been postulated that PrP(C) may be involved in one or more of the following: neurotransmitter metabolism, cell adhesion, signal transduction, copper metabolism, antioxidant activity or programmed cell death. Despite suggestive results supporting each of these mechanisms, the physiological function(s) of PrP(C) is not known. To investigate whether PrP(C) can prevent apoptotic cell death in prion diseases, we established the cell lines stably expressing PrP(C) from PrP knockout (PrP(-/-)) neuronal cells and examined the role of PrP(C) under apoptosis and/or serum-deprived condition. We found that PrP(-/-) cells were vulnerable to apoptotic cell death and that this vulnerability was rescued by the expression of PrP(C). The expression levels of apoptosis-related proteins including p53, Bax, caspase-3, poly(ADP-ribose) polymerase (PARP) and cytochrome c were significantly increased in PrP(-/-) cells. In addition, Ca(2+) levels of mitochondria were increased, whereas mitochondrial membrane potentials were decreased in PrP(-/-) cells. These results strongly suggest that PrP(C) may play a central role as an effective anti-apoptotic protein through caspase-dependent apoptotic pathways in mitochondria, supporting the concept that disruption of PrP(C) and consequent reduction of anti-apoptotic capacity of PrP(C) may be one of the pathogenic mechanisms of prion diseases.

Animals↗
PubMed1997

Mitochondrial dysfunction and calcium perturbation induced by traumatic brain injury.

Traumatic brain injury (TBI) is associated with primary and secondary injury. A thorough understanding of secondary injury will help to develop effective treatments and improve patient outcome. In this study, the GM model of controlled cortical impact injury (CCII) of Lighthall (1988) was used with modification to induce lateral TBI in rats. Forebrain mitochondria isolated from ipsilateral (IH) and contralateral (CH) hemispheres to impact showed a distinct difference. With glutamate + malate as substrates, mitochondria from the IH showed a significant decrease in State 3 respiratory rates, respiratory control indices (RCI), and P/O ratios. This decrease occurred as early as 1 h and persisted for at least 14 days following TBI. The State 3 respiratory rates, RCI, and P/O ratios could be restored to sham values by the addition of EGTA to the assay mixture. A significant amount of Ca2+ was found to be adsorbed to the mitochondria of both the IH and the CH with higher values seen in the IH. The rate of energy-linked Ca2+ transport in the IH was significantly decreased at 6 and 12 h. These data indicate that CCII-induced TBI perturbs cellular Ca2+ homeostasis and results in excessive Ca2+ adsorption to the mitochondrial membrane, which subsequently inhibits the respiratory chain-linked electron transfer and energy transduction.

Animals↗
PubMed2001

Clinical and laboratory signs of mitochondrial dysfunction secondary to nucleoside analogue antiretroviral therapy are reversible.

During 27 months treatment with 400 mg didanosine and 80 mg stavudine daily but no protease inhibitor therapy, a 50-y-old HIV-positive woman gradually lost 13 kg in weight, her arms, legs and buttocks decreased in volume and she experienced fatigue and polyneuropathy. Laboratory tests showed slight increases in plasma lactate and liver enzyme levels. Eighteen months after withdrawal of antiretroviral drug, the patient was free of fatigue and polyneuropathy and had regained 7 kg in weight as well as most of the volume of her arms, legs and buttocks.

Anti-HIV Agents↗
PubMed2003

Apoptosis induced by cryo-injury in human colorectal cancer cells is associated with mitochondrial dysfunction.

Cryotherapy, a method of in situ ablation, is used in the treatment of colorectal liver metastases with variable results. During the treatment, the central area of treated tumor undergoes necrotic destruction by lethal cryo-injury; however, the cellular response of tumor exposed to sublethal cryo-injury at the peripheral zone is unclear. In our study, we have identified the induction of apoptosis by cryo-injury at -10 degrees C in 4 colorectal cancer cell lines (HT29, HCT116, KM12C and KM12SM). The apoptosis was characterized by chromatin condensation, transferase-mediated dUTP nick end-labeling (TUNEL) staining, proteolytic cleavage of poly(ADP-ribose) polymerase (PARP) and cytokeratin 18, and activation of caspase-3. The occurrence and intensity of cryo-induced apoptosis did not correlate with the functional status of p53 in the cell lines studied. The expression of anti-apoptotic proteins (Bcl-2, Bcl-X(L)) and pro-apoptotic proteins (Bax, Bcl-X(S), Bad, and Bak) in response to cryo-injury varied in this cell line panel. The basal level of Bcl-2/Bax protein ratio correlated inversely to the apoptotic rate. We further demonstrated that Bax level decreased in cytosol and increased in mitochondria, followed by a loss of mitochondrial membrane potential after cryo-injury in HT29 cells. These findings indicate that cryo-injury induces apoptosis in colorectal cancer cells via disruption of mitochondrial integrity. The cryo-induced apoptosis was also identified in a nude mouse tumor xenograft model. Our elucidation of the apoptosis pathway induced by cryo-injury implies that synergistic combination of cryosurgery with pharmacological agents that augment of apoptosis induction may have clinical relevance in treating colorectal liver metastasis.

Animals↗
PubMed1999

Nitric oxide suppresses apoptosis via interrupting caspase activation and mitochondrial dysfunction in cultured hepatocytes.

Nitric oxide (NO) is a potent inhibitor of apoptosis in many cell types, including hepatocytes. We and others have described NO-dependent decreases in caspase activity in cells undergoing apoptosis. However, previous work has not determined whether NO disrupts the proteolytic processing and thus the activation of pro-caspases. Here we report that NO suppresses proteolytic processing and activation of multiple pro-caspases in intact cells, including caspase-3 and caspase-8. We found that both exogenous NO as well as endogenously produced NO via adenoviral inducible NO synthase gene transfer protected hepatocytes from tumor necrosid factor (TNF) alpha plus actinomycin D (TNFalpha/ActD)-induced apoptosis. Affinity labeling with biotin-VAD-fmk of all active caspase species in TNFalpha-mediated apoptosis identified four newly labeled spots (activated caspases) present exclusively in TNFalpha/ActD-treated cells. Both NO and the caspase inhibitor, Ac-DEVD-CHO, prevented the appearance of the four newly labeled spots or active caspases. Immunoanalysis of affinity labeled caspases demonstrated that caspase-3 was the major effector caspase. Western blot analysis also identified the activation of caspase-8 in the TNFalpha/ActD-treated cells, and the activation was suppressed by NO. Furthermore, NO inhibited several other events associated with caspase activation in cells, including release of cytochrome c from mitochondria, decrease in mitochondrial transmembrane potential, and cleavage of poly(ADP-ribose) polymerase in TNFalpha/ActD-treated cells. These findings indicate the involvement of multiple caspases in TNFalpha-mediated apoptosis in hepatocytes and establish the capacity of NO to inhibit not only active caspases but also caspase activation.

Animals↗