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Results for “mitochondrial dysfunction”

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PubMed1981

Mechanism of chlorpromazine-induced arrhythmia -- arrhythmia and mitochondrial dysfunction.

In this study, we investigated the mechanism of the arrhythmogenic action of chlorpromazine (CPZ). Thirty-two anesthetized mongrel dogs were used. In each, the chest was opened and a stimulating electrode was attached to the apex of the left ventricle and the ventricular multiple response threshold (VMRT) was measured. The carotid artery was cannulated to measure aortic pressure. The dogs were divided into four groups, and the time course of VMRT, blood pressure, and heart rate were determined. All groups were placed under observation for 30 min after CPZ infusion. In the control group, only saline (2ml/kg) was infused; CPZ group: CPZ (Img/kg) was infused 10 min after saline (2ml/kg) infusion; CoQ10 group: Coenzyme Q10 (CoQ10) (5mg/kg) was infused 10 min before CPZ (Img/kg) infusion; FAD group: Flavin-adenine-dinucleotide (FAD) (2mg/kg) was infused 10 min before CPZ (Img/kg) infusion. In each group, myocardial mitochondria were prepared 30 min after CPZ infusion. The mitochondrial functions, respiratory control index, AdP/O, State III rate of oxygen consumption, and activities of two segments of the electron-transport chain (NADH leads to CoQ leads to cyt.c and cyt.c leads to cyt.a,a3 leads to O2) were measured separately. Ca++--binding activity of the mitochondria was also determined. CPZ administration decreased VMRT and blood pressure, and caused mitochondrial dysfunction which derived from a disturbance in the first segment of the electron transport chain. Decreased Ca++--binding activity was observed when mitochondrial function was disturbed. CoQ10 prevented significantly the decrease in VMRT and the disturbance of mitochondrial function induced by CPZ, but did not prevent the hypotensive effect of CPZ. FAD prevented not only the decrease in VMRT and the disturbance of mitochondrial function, but also the hypotensive effect of CPZ. These results suggest that the decrease in VMRT is closely related to mitochondrial dysfunction induced by CPZ. Moreover, it is suggested that the arrhythmogenic effect of CPZ is derived from the decreased mitochondrial Ca++--binding activity.

Animals↗

PubMed2001

Mitochondrial dysfunction associated with a mutation in the Notch3 gene in a CADASIL family.

BACKGROUND: Cerebral autosomal arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is characterized by recurrent subcortical ischemic strokes and dementia caused by mutations in the Notch3 gene. In Drosophila melanogaster, Notch signaling has a pleiotropic effect, affecting most tissues of the organism during development. OBJECTIVE: To characterize a potential mitochondrial dysfunction associated with mutations in the Notch3 gene. METHODS: Biochemical, histochemical, molecular, and genetic analyses were performed on muscle biopsy specimens and fibroblasts obtained from patients of a Spanish family with CADASIL. Additional biochemical and molecular analyses of the N(55e11) mutant of D. melanogaster were performed. RESULTS: In muscle biopsy specimens, a significant decrease was found in the activity of complex I (NADH [reduced form of nicotinamide adenine dinucleotide] dehydrogenase), and in one patient, histochemical analysis showed the presence of ragged-red fibers with abnormal cytochrome c oxidase staining. Reduced fibroblast activity of complex V (ATP synthase) was found. Supporting data on patients with CADASIL, it was found that the mutation N(55e11) in Drosophila decreases the activity of mitochondrial respiratory complexes I and V. CONCLUSIONS: Mitochondrial respiratory chain activity responds, directly or indirectly, to the Notch signaling pathway. Mitochondrial dysfunction in patients with CADASIL may be an epiphenomenon, but results of this study suggest that the pathophysiology of the disease could include a defect in oxidative phosphorylation.

Adult↗

PubMed2002

Neurolathyrism: mitochondrial dysfunction in excitotoxicity mediated by L-beta-oxalyl aminoalanine.

beta-N-Oxalyl amino-L-alanine (L-BOAA); synonym beta-N-oxalyl-alpha,beta-diaminopropionic acid (beta-ODAP) is a naturally occurring non-protein amino acid present in the chickling pea from the plant Lathyrus sativus grown in drought prone areas. Ingestion of L-BOAA as a staple diet results in a progressive neurodegenerative condition, neurolathyrism, a form of motor neuron disease which affects the upper motor neurons and anterior horn cells of the lumbar spinal cord. L-BOAA is an excitatory acid and acts as an agonist at the AMPA receptor. One of the primary effects of L-BOAA toxicity is the inhibition of mitochondrial complex I selectively in the motor cortex and lumbar spinal cord. Recent evidence has suggested that the mitochondrial dysfunction is a consequence of oxidation protein thiol groups as a result of generation of reactive oxygen species. Mitochondrial complex I is highly to vulnerable to inactivation through oxidation of vital sulfhydryl groups. Thiol antioxidants such as alpha-liopic acid offer a method of protecting mitochondrial function. A common mechanism involving oxidation of protein thiol groups may underlie neurodegeneration occurring through mitochondrial dysfunction induced by excitatory amino acid.

Amino Acids, Diamino↗

PubMed2005

Proteomics analysis provides insight into caloric restriction mediated oxidation and expression of brain proteins associated with age-related impaired cellular processes: Mitochondrial dysfunction, glutamate dysregulation and impaired protein synthesis.

Age-related impairment of functionality of the central nervous system (CNS) is associated with increased susceptibility to develop many neurodegenerative diseases. Increased oxidative stress in the CNS of aged animals is manifested by increased protein oxidation, which is believed to contribute to the age-related learning and memory deficits. Glutamate dysregulation, mitochondrial dysfunction and impaired protein synthesis are observed in aged brains, along with increased protein oxidation. Interestingly, all of these age-related cellular alterations can be improved by caloric restriction (CR), which can also improve the plasticity and recovery of the CNS. Although the beneficial effects of CR on brains are well established, the mechanism(s) of its action remains unclear. In order to gain insight into the mechanism of CR in the brain, we located the brain regions that are benefited the most from reduced oxidative stress by CR. Along with other brain regions, striatum (ST) showed significantly decreased bulk protein carbonyl levels and hippocampus (HP) showed decreased bulk protein 3-nitrotyrosine (3-NT) levels in CR aged rats when compared to those of age matched controls. To determine which proteins were oxidatively modified in these brain regions, we used parallel proteomics approach to identify the proteins that are altered in oxidation and expression. The specific carbonyl levels of pyruvate kinase M2 (PKM2), alpha-enolase (ENO1), inositol monophosphatase (INSP1), and F1-ATPase Chain B (ATP-F1B) were significantly decreased in ST of aged CR rats. In contrast, the expression levels of phosphoglycerate kinase 1 (PKG1), inosine monophosphate cyclohydrolase (IMPCH) and F1-ATPase Chain A (ATP-F1A) were significantly increased in the ST of CR rats. In the hippocampus of CR rats, the specific 3-NT levels of malate dehydrogenase (MDH), phosphoglycerate kinase 1 (PKG1) and 14-3-3 zeta protein were significantly decreased and expression levels of DLP1 splice variant 1 (DLP1), mitochondrial aconitase (ACO2), dihydrolipoamide dehydrogenase (DLDH), neuroprotective peptide H3 (NPH3), and eukaryotic translation initiation factor 5A (eIF-5A) are increased. Moreover, an unnamed protein product (UNP1) with similar sequence to initiation factor 2 (IF-2) was decreased in the HP of CR rats. Our data support the hypothesis that CR induces a mild metabolic stress response by increasing the production of neurotrophic proteins, therefore, priming neurons against apoptosis. Moreover, our study shows that the improvement of glutamate dysregulation, mitochondrial dysfunction and protein synthesis by CR is, at least partially, due to the CR-mediated alteration of the oxidation or the expression of PKM2, ENO1, INSP1, ATP-F1B, PKG1, IMPCH, ATP-F1A MDH, PKG1 and 14-3-3 zeta protein, DLP1, ACO2, DLDH, NPH3, eIF-5A and UNP1. This study provides valuable insights into the mechanisms of the beneficial factors on brain aging by CR.

Aging↗

PubMed1999

The antioxidant EPC-K1 attenuates NO-induced mitochondrial dysfunction, lipid peroxidation and apoptosis in cerebellar granule cells.

In this study we investigated the effects of nitric oxide (NO) on cultured cerebellar granule cells. Exposure to NO donors, S-nitrosoglutathione (GSNO; 250 microM) or sodium nitroprusside (SNP; 500 microM), triggered apoptosis in immature cultures of cerebellar granule cells, which was characterized by chromatin condensation, nuclei fragmentation, and DNA laddering. Exposure of cerebellar granule cells to NO donors led to a decrease in the mitochondrial transmembrane potential and intracellular ATP content, which suggested that NO treatment caused mitochondrial dysfunction. NO treatment also induced oxidative stress in cerebellar granule cells as measured by thiobarbituric acid (TBA) assay. Pretreating cells with L-ascorbic acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-2H -1-benzopyran-6-yl-hydrogen phosphate] potassium salt (EPC-K1), a novel antioxidant, attenuated NO-induced mitochondrial dysfunction and oxidative stress to some extent, and prevented the cells from apoptosis. The results of the present investigation suggest that a superoxide/peroxynitrite-mediated oxidative stress may be an important pathway leading to NO-associated neuronal damage. Pretreating cells with the antioxidant EPC-K1 attenuated NO-induced neurotoxicity by scavenging superoxide/peroxynitrite and/or its breakdown products.

Animals↗

PubMed2003

Tea catechins protect against lead-induced ROS formation, mitochondrial dysfunction, and calcium dysregulation in PC12 cells.

Recent studies have shown that lead causes oxidative stress by inducing the generation of reactive oxygen species (ROS) and reducing the antioxidant defense system of cells, which suggests that antioxidants may play an important role in the treatment of lead poisoning. The present study was designed to elucidate whether tea catechins had any protective effects on altered oxidative stress parameter in PC12 cells exposed to lead. The experimental results showed that lead decreased PC12 cell viability and induced a rapid elevation of [Ca(2+)](i), which was followed by an accumulation of ROS and a decrease of mitochondrial membrane potential (MMP). Treatment by tea catechins significantly increased cell viability, decreased intracellular Ca(2+) levels and ROS formation, and improved MMP in PC12 cells exposed to lead. The galloylated catechins showed a greater effect on ROS formation and mitochondrial dysfunction than that of nongalloylated catechins, which was similar to the result of their scavenging ability on free radical. In view of the time course of ROS formation and mitochondrial dysfunction and their correlation, our results also suggested that the beneficial effects of tea catechins on MMP are related, at least in part, to its ability to scavenge ROS in PC12 cells exposed to 100 microM Pb(2+). The present results suggest that tea catechins supplementation may play a role for modulating oxidative stress in PC12 cells exposed to lead.

Animals↗

PubMed1999

Altered calcium homeostasis and mitochondrial dysfunction in cortical synaptic compartments of presenilin-1 mutant mice.

Alzheimer's disease is characterized by amyloid beta-peptide deposition, synapse loss, and neuronal death, which are correlated with cognitive impairments. Mutations in the presenilin-1 gene on chromosome 14 are causally linked to many cases of early-onset inherited Alzheimer's disease. We report that synaptosomes prepared from transgenic mice harboring presenilin-1 mutations exhibit enhanced elevations of cytoplasmic calcium levels following exposure to depolarizing agents, amyloid beta-peptide, and a mitochondrial toxin compared with synaptosomes from nontransgenic mice and mice overexpressing wild-type presenilin-1. Mitochondrial dysfunction and caspase activation following exposures to amyloid beta-peptide and metabolic insults were exacerbated in synaptosomes from presenilin-1 mutant mice. Agents that buffer cytoplasmic calcium or that prevent calcium release from the endoplasmic reticulum protected synaptosomes against the adverse effect of presenilin-1 mutations on mitochondrial function. Abnormal synaptic calcium homeostasis and mitochondrial dysfunction may contribute to the pathogenic mechanism of presenilin-1 mutations.

Alzheimer Disease↗

PubMed2005

Insulin resistance in the HIV-infected population: the potential role of mitochondrial dysfunction.

Insulin resistance is accepted as the underlying fundamental defect that predates and ultimately leads to the development of type 2 (adult onset) diabetes mellitus in the general non-human immunodeficiency virus (HIV)-infected population. Insulin resistance is also a major component of the metabolic syndrome that, in association with other factors such as hypertension, hypercholesterolemia, and central obesity, defines a pre-diabetic atherogenic state that leads to adverse cardiovascular events. Growing evidence now suggests that mitochondrial dysfunction in skeletal muscle may be the mechanism whereby insulin resistance is induced. The prevalence of insulin resistance, glucose intolerance, and diabetes in the HIV-infected population has dramatically increased following the common use of highly active antiretroviral therapy (HAART). The development of insulin resistance in the HIV-infected population is likely to be multifactorial reflecting genetic predisposition, direct and indirect effects of both the protease inhibitor (PI) and nucleoside reverse transcriptase inhibitor (NRTI) class of antiretroviral therapy, and a possible contribution from chronic inflammatory changes induced by HIV. Indirect effects of antiretroviral therapy on insulin resistance may be mediated through both the visceral adiposity and peripheral fat depletion components of lipodystrophy as well as through fatty infiltration in liver and muscle. Based on current knowledge, mitochondrial dysfunction can be hypothesized to play a key role in each of these components.

Acquired Immunodeficiency Syndrome↗

PubMed2003

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) promotes mitochondrial dysfunction and apoptosis induced by 7-hydroxystaurosporine and mitogen-activated protein kinase kinase inhibitors in human leukemia cells that ectopically express Bcl-2 and Bcl-xL.

Previous studies have demonstrated that cotreatment with mitogen activated-protein kinase kinase (MEK) 1/2 inhibitors (e.g., PD184352) and the checkpoint abrogator 7-hydroxystaurosporine (UCN-01) dramatically induces apoptosis in a variety of human leukemia and multiple myeloma cell types. The purpose of this study was to evaluate the roles of Bcl-2 family members and the relative contribution of the intrinsic mitochondrial versus the extrinsic receptor-related apoptotic pathways to MEK inhibitors/UCN-01-induced leukemic cell death. Cotreatment of U937 cells with PD184352 and UCN-01 resulted in the activation of procaspase-3, -9, and -8 as well as Bid cleavage. PD184352/UCN-01-induced mitochondrial dysfunction and apoptosis were both substantially attenuated in cells ectopically expressing Bcl-2, an N-terminal phosphorylation loop-deleted mutant Bcl-2, or Bcl-xL, but not in cells expressing dominant-negative (DN) caspase-8, cytokine response modifier A (cowpox virus-encoded antiapoptotic protein), or DN Fas-associated death domain. Coadministration of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) or TNF-alpha substantially increased MEK inhibitors (e.g., PD184352 or U0126)/UCN-01-induced mitochondrial dysfunction, activation of procaspase-8 and Bid, and apoptosis in Bcl-2- and Bcl-xL-overexpressing cells but not in those in which the extrinsic pathway was interrupted. Together, these findings suggest that the MEK inhibitors/UCN-01 regimen primarily induces leukemic cell apoptosis by engaging the intrinsic, mitochondrial apoptotic pathway and that resistance to these events conferred by increased expression of certain antiapoptotic Bcl-2 family members can be overcome, at least in part, by coadministration of TRAIL and other agents that activate the extrinsic apoptotic cascade.

Apoptosis↗

PubMed2005

Physiologic progesterone reduces mitochondrial dysfunction and hippocampal cell loss after traumatic brain injury in female rats.

Growing literature suggests important sex-based differences in outcome following traumatic brain injury (TBI) in animals and humans. Progesterone has emerged as a key hormone involved in many potential neuroprotective pathways after acute brain injury and may be responsible for some of these differences. Many studies have utilized supraphysiologic levels of post-traumatic progesterone to reverse pathologic processes after TBI, but few studies have focused on the role of endogenous physiologic levels of progesterone in neuroprotection. We hypothesized that progesterone at physiologic serum levels would be neuroprotective in female rats after TBI and that progesterone would reverse early mitochondrial dysfunction seen in this model. Female, Sprague-Dawley rats were ovariectomized and implanted with silastic capsules containing either low or high physiologic range progesterone at 7 days prior to TBI. Control rats received ovariectomy with implants containing no hormone. Rats underwent controlled cortical impact to the left parietotemporal cortex and were evaluated for evidence of early mitochondrial dysfunction (1 h) and delayed hippocampal neuronal injury and cortical tissue loss (7 days) after injury. Progesterone in the low physiologic range reversed the early postinjury alterations seen in mitochondrial respiration and reduced hippocampal neuronal loss in both the CA1 and CA3 subfields. Progesterone in the high physiologic range had a more limited pattern of hippocampal neuronal preservation in the CA3 region only. Neither progesterone dose significantly reduced cortical tissue loss. These findings have implications in understanding the sex-based differences in outcome following acute brain injury.

Animals↗

PubMed1995

Nitric oxide mediates mitochondrial dysfunction in hepatoma cells induced by non-activated Kupffer cells: evidence implicating ICAM-1-dependent process.

The metabolic changes in a rat hepatoma cell line, AH70 cells, after co-culture with rat Kupffer cells (KC) were visualized and analysed using a fluorescence microscope equipped with a silicon intensified target camera and a laser scanning confocal microscopic system. Kupffer cells were isolated from male Wistar rats, and cultured without any stimuli. The non-activated KC reduced the mitochondrial energization in the cocultured AH70 cells within 2 h, which was indicated by decreased rhodamine 123 (Rh123) fluorescence. Either NG-monomethyl-L-arginine or dexamethasone significantly attenuated the KC-induced mitochondrial dysfunction in AH70 cells, suggesting the involvement of nitric oxide (NO) derived from inducible-type nitric oxide synthase (iNOS). Administration of monoclonal antibody (mAb) directed against rat ICAM-1 also prevented the decrease in Rh123 fluorescence. Electron microscopy revealed that the membrane-to-membrane attachment between KC and AH70 cells occurred within 2 h. A laser scanning confocal microscopic observation using mAb against ICAM-1 presented that the ICAM-1 expression on AH70 cells and KC increased after the co-culture. It is therefore concluded that the KC-mediated mitochondrial dysfunction of hepatoma cells largely depends on NO production by iNOS. Furthermore, the present study supports a scenario that the NO production and release from KC is triggered by the close contact with hepatoma cells through adhesion molecules such as ICAM-1.

Animals↗

PubMed2003

Role of protein aggregation in mitochondrial dysfunction and neurodegeneration in Alzheimer's and Parkinson's diseases.

Abnormal interactions and misfolding of synaptic proteins in the nervous system are being extensively explored as important pathogenic events resulting in neurodegeneration in various neurological disorders. These include Alzheimer's disease (AD), Parkinson's disease (PD), and dementia with Lewy bodies (DLB). In AD, misfolded amyloid beta peptide 1-42 (Abeta), a proteolytic product of amyloid precursor protein metabolism, accumulates in the neuronal endoplasmic reticulum and extracellularly as plaques. In contrast, in PD and DLB cases there is abnormal accumulation of alpha-synuclein in neuronal cell bodies, axons, and synapses. Furthermore, in DLB, Abeta 1-42 may promote alpha-synuclein accumulation and neurodegeneration. The central event leading to synaptic and neuronal loss in these diseases is not completely clear yet; however, recent advances in the field suggest that nerve damage might result from the conversion of nontoxic monomers to toxic oligomers and protofibrils. The mechanisms by which misfolded Abeta peptide and alpha-synuclein might lead to synapse loss are currently under investigation. Several lines of evidence support the possibility that Abeta peptide and alpha-synuclein might interact to cause mitochondrial and plasma membrane damage upon translocation of protofibrils to the membranes. Accumulation of Abeta and alpha-synuclein oligomers in the mitochondrial membrane might result in the release of cytochrome C with the subsequent activation of the apoptosis cascade. Conversely, the oxidative stress and mitochondrial dysfunction associated with AD and PD may also lead to increased membrane permeability and cytochrome C release, which promotes Abeta and alpha-synuclein oligomerization and neurodegeneration. Together, these studies suggest that the translocation of misfolded proteins to the mitochondrial membrane might play an important role in either triggering or perpetuating neurodegeneration. The insights obtained from the characterization of this process may be applied to the role of mitochondrial dysfunction in other neurodegenerative disorders, including AD. New evidence may also provide a rationale for the mitochondrial membrane as a target for therapy in a variety of neurodegenerative diseases.

Alzheimer Disease↗

PubMed2003

Maternal treatment with alpha-phenyl-N-tert-butylnitrone attenuates secondary mitochondrial dysfunction after transient intrauterine asphyxia in the fetal rat brain.

Recirculation following 30 min of ischemia due to transient uterine artery occlusion in pregnant rats at 20 days gestation has previously been found to cause partial recovery and secondary deterioration of the cellular bioenergetic state in the fetal brain, the subsequent damage being ameliorated by a free radical spin trap agent, alpha-phenyl-N-TERT-butylnitrone (PBN). Our objective was to assess whether the secondary deterioration of the cellular bioenergetic state is due to mitochondrial dysfunction and to study whether PBN acts to prevent secondary damage to mitochondria in the fetal rat brain. Fetal neocortical tissues were sampled after 30 min of intrauterine ischemia and after 1, 2 or 4 h of recirculation. PBN or vehicle was given 1 h after recirculation. Homogenates were prepared, and ADP-stimulated, nonstimulated and uncoupled respiratory rates were measured polarographically. Ischemia was associated with a decrease in ADP-stimulated and uncoupled respiratory rates, with a marked fall in the respiratory control ratio, defined as ADP-stimulated divided by nonstimulated respiration (p < 0.01). Recirculation (1 h) brought about partial recovery, but continued reflow (2 and 4 h) was associated with a secondary deterioration of respiratory functions (p < 0.01). The secondary deterioration was prevented by PBN (p < 0.05). The results demonstrate that the secondary deterioration of the cellular bioenergetic state in ischemia-reperfusion is due to secondary mitochondrial dysfunction and that this deterioration may be induced by oxygen-derived free radicals in the immature fetal brain.

Adenosine Diphosphate↗

PubMed2001

Electron transport chain defect and inefficient respiration may underlie pulmonary hypertension syndrome (ascites)-associated mitochondrial dysfunction in broilers.

By using a series of chemical inhibitors of mitochondrial respiration, a site-specific defect in the electron transport chain was identified in mitochondria obtained from broilers with pulmonary hypertension syndrome (PHS; ascites). Located at the succinate:ubiquinone oxido-reductase (Complex II:CoQ) interface, this defect would allow electrons to leak from the respiratory chain and consume oxygen by forming reactive oxygen species at a greater rate than in control mitochondria. Lower levels of the primary antioxidants, alpha- and beta-tocopherol, and glutathione (GSH) in PHS mitochondria confirmed the presence of oxidative stress. Respiration studies of PHS liver mitochondria also revealed disease-associated decreases in the respiratory control ratio (RCR, an index of electron transport chain coupling). Differences in the RCR as well as the adenosine diphosphate (ADP) to O ratio (an index of oxidative phosphorylation) between control and PHS mitochondria were accentuated by sequential additions of ADP to isolated mitochondria. In a second experiment, similar improvements in functional indices following sequential additions of ADP and responses to respiratory chain inhibitors were observed in liver mitochondria isolated from Single Comb White Leghorn (SCWL) males (resistant to PHS) similar to that observed in control broiler mitochondria in Experiment 1. The combined results indicate the presence of a site-specific defect at either Complex II, ubiquinone, or both in liver mitochondria obtained from broilers with PHS that may be responsible for the oxidative stress and mitochondrial dysfunction observed in this costly metabolic disease.

Adenosine Diphosphate↗

PubMed2000

Distinct stages of cytochrome c release from mitochondria: evidence for a feedback amplification loop linking caspase activation to mitochondrial dysfunction in genotoxic stress induced apoptosis.

Cytochrome c (cyto c) release from mitochondria is a critical event in apoptosis. By investigating the ordering of molecular events during genotoxic stress-induced apoptosis, we found that ionizing radiation (IR) and etoposide induced the release of cyto c from mitochondria in two distinct stages. The early release of low levels of cyto c into the cytosol preceded the activation of caspase 9 and 3, but had no effect on ATP levels or mitochrondrial transmembrane potential (Deltapsim). In contrast, the late stage cyto c release resulted in a drastic loss of mitochondrial cyto c and was associated with reduction of ATP levels and Deltapsim. Moreover, caspases contributed to the late cyto c release since the caspase inhibitor zVAD prevented only the late but not the early-stage cyto c release. Recombinant caspase 3 induced cyto c release from isolated mitochondria in the absence of cytosolic factors. Bcl-2 but not Bid was cleaved during apoptosis after caspase activation. This suggests that Bcl-2 cleavage might contribute to the late cyto c release, which results in mitochondrial dysfunction manifested by the decrease of ATP and Deltapsim. zVAD prevented the reduction of ATP, Deltapsim, and nuclear condensation when added up to 8 h after IR, at the time the caspases were highly activated but when the majority of cyto c was still maintained in the mitochondria. These findings link the feedback loop control of caspase-induced cyto c release with mitochondrial dysfunction manifested by ATP and Deltapsim decline.

Apoptosis↗

PubMed2004

Benzo(a)pyrene-induced apoptotic death of mouse hepatoma Hepa1c1c7 cells via activation of intrinsic caspase cascade and mitochondrial dysfunction.

Benzo(a)pyrene (BaP), a potent carcinogen, has been shown to induce apoptosis via activation of caspase-3. However, the upstream of caspase-3 and other apoptosis signaling remain to be elusive. Herein, we demonstrated that treatment of Hepa1c1c7 cells with BaP increased the transcriptional expression of aryl hydrocarbon nuclear transporter and cytochrome p450 1A1 in a time and dose-dependent manner but did not aromatic hydrocarbon receptor. Also, the catalytic activation of caspase-3 and caspase-9 was induced whereas that of caspase-3 and caspase-9 was not by the addition of BaP. BaP also induced the mitochondrial dysfunction, including transition of mitochondria membrane potential and cytosolic release of cytochrome c. Furthermore, a decrease in the expression of Bcl-2 to Bax ratio and phosphorylation of p53(Ser 15) were observed in BaP-treated cells. Taken together, these results demonstrated that BaP-induced apoptosis of Hepa1c1c7 cells via activation of intrinsic caspase pathway including caspase-3, caspase-9, with mitochondrial dysfunction and p53 activation.

Animals↗

PubMed2003

Flavopiridol potently induces small cell lung cancer apoptosis during S phase in a manner that involves early mitochondrial dysfunction.

PURPOSE: Accumulating evidence indicates that small cell lung cancer (SCLC) is defective in many of the regulatory mechanisms that control cell cycle progression. The purpose of this study was to determine the effects of flavopiridol, a pan-cyclin-dependent kinase inhibitor, on growth and apoptosis of SCLC cell lines. EXPERIMENTAL DESIGN: Cell growth was monitored using 3-(4,5dimethylthiazol-2yl)-2,5-diphenyl-tetrazolium bromide (MTT) and clonogenic assays. Induction of apoptosis was assessed using multiple assays, including flow cytometric determination of DNA content and mitochondrial membrane potential, terminal deoxynucleotide transferase-mediated dUTP nick end labeling (TUNEL), and Western blot analysis of procaspase 3 and poly(ADP-ribose) polymerase cleavage. RESULTS: Flavopiridol induced growth inhibition and cytotoxicity in multiple SCLC cell lines, with an IC(50) of 50-100 nM and an LD(50) of 150-200 nM in 72-h MTT assays. The cytotoxicity seen in the MTT assay proved to be apoptosis by several criteria. Interestingly, inhibition of caspase activation with the caspase inhibitor Boc-Asp(OMe)-CH(2)F reduced TUNEL labeling by 40% but did not have any effect on the loss of mitochondrial membrane potential (detected as early as 4 h after drug exposure) or cytotoxicity in MTT assays. These results suggest that the primary event in flavopiridol-induced apoptosis involves induction of mitochondrial dysfunction. Cells synchronized with aphidicolin at the G(1)-S border and treated with flavopiridol during S phase showed a marked increase in apoptosis compared with an asynchronous population or a population treated during G(2)-M. Despite the increased apoptosis, a significant proportion of synchronized cells proceeded through S, G(2)-M, and into G(1) phase in the presence of flavopiridol, demonstrating that a high-grade cell cycle arrest is not required for apoptosis. Cells synchronized at the G(1)-S border treated with a short exposure to flavopiridol also showed more than a 10-fold decrease in clonogenicity compared with asynchronous cells treated identically. CONCLUSIONS: Taken together, these data demonstrate that flavopiridol potently and selectively induces SCLC apoptosis preferentially during S phase, in a manner that involves early mitochondrial dysfunction without a requirement for a high-grade block to cell cycle progression. Furthermore, clonogenicity data suggests that prior S phase synchronization could be a highly effective way of enhancing the efficacy of bolus or short infusions of flavopiridol in the clinical setting.

Aphidicolin↗

PubMed1997

HIV-1 protein Vpr causes gross mitochondrial dysfunction in the yeast Saccharomyces cerevisiae.

The biological effects of the HIV-1 accessory protein, Vpr, have been studied in yeast expression systems. In our previous study [1], employing the pCUP1-vpr copper-inducible expression cassette, Vpr was shown to cause growth arrest and structural defects. In this study yeast constitutively expressing vpr, through elevated copy number and/or elevated transcription levels, displayed no growth arrest in fermentative growth conditions while Vpr was produced at much lower levels than in the inducible expression system. However, such cells were respiratory deficient and unable to utilise ethanol or glycerol as the sole carbon source. They exhibited gross mitochondrial dysfunction displayed in the loss of respiratory chain complex I, II, III, IV and citrate synthase activities. The effects on mitochondria required a C-terminal domain of Vpr that contains a conserved amino acid sequence motif HFRIGCRHSRIG. These results suggest that the widely observed phenomenon of 'Vpr-induced growth arrest' in human cells could be due to mitochondrial dysfunction.

Gene Products, vpr↗