Disruption Of Mitochondrial Metabolism Research Articles

Cracking the cytotoxicity code: apoptotic induction of 10-acetylirciformonin B is mediated through ROS generation and mitochondrial dysfunction.

A marine furanoterpenoid derivative, 10-acetylirciformonin B (10AB), was found to inhibit the proliferation of leukemia, hepatoma, and colon cancer cell lines, with selective and significant potency against leukemia cells. It induced DNA damage and apoptosis in leukemia HL 60 cells. To fully understand the mechanism behind the 10AB apoptotic induction against HL 60 cells, we extended our previous findings and further explored the precise molecular targets of 10AB. We found that the use of 10AB increased apoptosis by 8.9%–87.6% and caused disruption of mitochondrial membrane potential (MMP) by 15.2%–95.2% in a dose-dependent manner, as demonstrated by annexin-V/PI and JC-1 staining assays, respectively. Moreover, our findings indicated that the pretreatment of HL 60 cells with N-acetyl-l-cysteine (NAC), a reactive oxygen species (ROS) scavenger, diminished MMP disruption and apoptosis induced by 10AB, suggesting that ROS overproduction plays a crucial rule in the cytotoxic activity of 10AB. The results of a cell-free system assay indicated that 10AB could act as a topoisomerase catalytic inhibitor through the inhibition of topoisomerase IIα. On the protein level, the expression of the anti-apoptotic proteins Bcl-xL and Bcl-2, caspase inhibitors XIAP and survivin, as well as hexokinase II were inhibited by the use of 10AB. On the other hand, the expression of the pro-apoptotic protein Bax was increased after 10AB treatment. Taken together, our results suggest that 10AB-induced apoptosis is mediated through the overproduction of ROS and the disruption of mitochondrial metabolism.

The disruption of mitochondrial metabolism and ion homeostasis in chicken hearts exposed to manganese

Exposure to high levels of manganese (Mn) can result in cardiotoxicity in animals. However, little is known about the effect of excess Mn on poultry hearts. The aim of this study was to investigate the effect of dietary Mn on chicken cardiac injuries and the possible mechanisms of this process. In the present study, 400 fifty-day-old Hy-line brown cocks were randomly divided into four groups, and were fed either a commercial diet (containing 100mg/kg Mn) or a Mn-supplemented diet containing 600mg/kg, 900mg/kg, or 1800mg/kg Mn for 30, 60 or 90 days, respectively. Next, we examined several biomarkers of cardiac injury, including biochemical blood serum analyses, electrocardiogram assays, histological analyses, ultra-structural assays and apoptosis assays. To investigate the possible mechanisms of Mn-induced cardiotoxicity, we examined the effect of MnCl2 on mitochondrial function and metal ion homeostasis. We found that subchronic MnCl2 exposure induced damage in chicken hearts. Further investigations indicated that possible mechanisms for Mn-induced chicken cardiac injury included the disruption of mitochondrial metabolism and the alteration of ion homeostasis.

Mitochondrial oxidative stress and the metabolic syndrome

The current epidemic of the metabolic syndrome in the developed world is largely due to overnutrition and lack of physical activity. However, the underlying causes by which chronic overnutrition interacts with genotype and physical inactivity to generate the metabolic syndrome phenotype are complex, and include multiple metabolic and physiological alterations. Mitochondrial oxidative stress has been suggested to contribute to the metabolic syndrome, but the mechanisms and significance are unclear. Here we review how disruption of mitochondrial metabolism and increased oxidative stress may occur during overnutrition coupled with limited physical activity. From this we suggest a unifying hypothesis to integrate what is known about mitochondrial involvement in the metabolic syndrome that points to testable hypotheses and novel therapeutic approaches.

Non-redox-active lipoate derivates disrupt cancer cell mitochondrial metabolism and are potent anticancer agents in vivo

We report the analysis of CPI-613, the first member of a large set of analogs of lipoic acid (lipoate) we have investigated as potential anticancer agents. CPI-613 strongly disrupts mitochondrial metabolism, with selectivity for tumor cells in culture. This mitochondrial disruption includes activation of the well-characterized, lipoate-responsive regulatory phosphorylation of the E1α pyruvate dehydrogenase (PDH) subunit. This phosphorylation inactivates flux of glycolysis-derived carbon through this enzyme complex and implicates the PDH regulatory kinases (PDKs) as a possible drug target. Supporting this hypothesis, RNAi knockdown of the PDK protein levels substantially attenuates CPI-613 cancer cell killing. In both cell culture and in vivo tumor environments, the observed strong mitochondrial metabolic disruption is expected to significantly compromise cell survival. Consistent with this prediction, CPI-613 disruption of tumor mitochondrial metabolism is followed by efficient commitment to cell death by multiple, apparently redundant pathways, including apoptosis, in all tested cancer cell lines. Further, CPI-613 shows strong antitumor activity in vivo against human non-small cell lung and pancreatic cancers in xenograft models with low side-effect toxicity.

Cells Silenced forSDHBExpression Display Characteristic Features of the Tumor Phenotype

Recently, enzymes of the tricarboxylic acid (TCA) cycle have emerged as novel tumor suppressors. In particular, mutations in the nuclear-encoded subunits of succinate dehydrogenase (SDHB, SDHC, and SDHD) cause paragangliomas and pheochromocytomas. Although the mechanism(s) by which disruption of mitochondrial metabolism leads to neoplasia is largely unknown, increasing evidence points to an activation of pseudohypoxia. In this study, we have shown that silencing of SDHB using DNA-based small interfering RNA resulted in major impairments in cellular proliferation, respiration, and a corresponding shift to glycolysis. The levels of reactive oxygen species, however, were unchanged. As expected, hypoxia-inducible factor-1 alpha (HIF-1 alpha) and HIF-2alpha were up-regulated in chronically silenced cells, suggesting that a pseudohypoxic state was attained. In addition, the c-Jun amino-terminal kinase and p38 kinase stress signaling proteins were hyperphosphorylated in SDHB-silenced cells. Microarray analysis showed that >400 genes were influenced (6-fold or more up-regulation or down-regulation) by silencing of SDHB, confirming the importance of the TCA cycle in cellular metabolism. Examples of dysregulated genes included those involved in proliferation, adhesion, and the hypoxia pathway. Of interest, SDHB-silenced cells had a greater capacity to adhere to extracellular matrix components, including fibronectin and laminin, than control cells, thus suggesting a possible mechanism of tumor initiation. Although transient silencing of the HIF-1 alpha transcription factor in SDHB-silenced cells had little effect on the expression of a subset of up-regulated genes, it partially reversed the adhesion phenotype to fibronectin, pointing to a potentially important role for HIF-1 in this process.

214. Altered placental gene expression following disruption of mitochondrial metabolism in mouse embryos

Metabolic changes in the preimplantation embryo are known to alter blastocyst viability. We used a mouse embryo model, inhibiting the malate-aspartate shuttle (MAS) which is required for energy metabolism in the early embryo, to examine the consequences on subsequent fetal and placental development, and placental gene expression. Mouse zygotes were obtained from C57BL6xCBA females after gonadotrophin stimulation. Following culture in G1.2 for 48h, 8-cell embryos were cultured to the blastocyst stage in either G2.2 control media (C), G2.2 media without pyruvate (-P), or -P with 0.5 mM aminooxyacetate, an inhibitor of MAS (-P+AOA). Blastocysts were transferred to day 4 pseudopregnant Swiss mice, and fetuses and placentas were harvested on day 18 of pregnancy. RNA was extracted from placentas for real time PCR expression analysis by ddCt relative to 18S. Expression of mitochondrial transcription factors (mTERF, mTFAM, Nrf-1, Nrf-2), glucose transporters (Glut1 and Glut3) and amino acid transporters (Slc38a2 and Slc38a4) were analysed. Following transfer, fetal development per implantation was significantly reduced when embryos were cultured in –P+AOA conditions (25.9%) relative to both C and –P conditions (57.5% and 68.8% respectively). Although placental weight did not differ between treatment groups, fetal weight was significantly reduced for –P and –P+AOA groups (P < 0.05), suggestive of altered placental function thus gene expression was examined. There were no differences in gene expression between placentas from C and –P conditions for any of the genes analysed. When compared with C placentas, -P+AOA conditions reduced the expression of Glut 1 (P = 0.055) and Glut 3 (P = 0.043). The –P+AOA placentas also tended to have significantly reduced expression of Glut 3 (P = 0.06) compared with placentas from –P conditions, but all other genes were expressed similarly. This data suggests that decreased glucose transport may be the cause of reduced fetal growth as a consequence of metabolic perturbations in the embryo.

6564 POSTER Frontline cytotoxic chemotherapy (CTx) for newly diagnosed non-small cell lung cancer (NSCLC) patients presenting with brain metastasis compared to whole brain radiotherapy (WBRT): result of a randomized pilot study

Exposure to high levels of manganese (Mn) can result in cardiotoxicity in animals. However, little is known about the effect of excess Mn on poultry hearts. The aim of this study was to investigate the effect of dietary Mn on chicken cardiac injuries and the possible mechanisms of this process. In the present study, 400 fifty-day-old Hy-line brown cocks were randomly divided into four groups, and were fed either a commercial diet (containing 100 mg/kg Mn) or a Mn-supplemented diet containing 600 mg/kg, 900 mg/kg, or 1800 mg/kg Mn for 30, 60 or 90 days, respectively. Next, we examined several biomarkers of cardiac injury, including biochemical blood serum analyses, electrocardiogram assays, histological analyses, ultra-structural assays and apoptosis assays. To investigate the possible mechanisms of Mn-induced cardiotoxicity, we examined the effect of MnCl2 on mitochondrial function and metal ion homeostasis. We found that subchronic MnCl2 exposure induced damage in chicken hearts. Further investigations indicated that possible mechanisms for Mn-induced chicken cardiac injury included the disruption of mitochondrial metabolism and the alteration of ion homeostasis.

Recurrent pancreatitis in mitochondrial cytopathy

Diabetes mellitus and exocrine insufficiency are the commonest pancreatic manifestations of mitochondrial diseases. In contrast, pancreatitis has rarely been described in mitochondrial syndromes. We report on a patient with Kearns-Sayre syndrome and recurrent episodes of acute pancreatitis for which no explanation could be found other than the associated mitochondrial dysfunction. Interestingly, pharmacological disruption of mitochondrial metabolism in various models as well as in patients can cause pancreatitis, further supporting this association. A diagnosis of pancreatitis should be considered in any patients with mitochondrial disease and recurrent abdominal pain.

Long-Term Effect of Acetyl-L-Carnitine for Antiretroviral Toxic Neuropathy

Background: Nucleoside analogue reverse transcriptase inhibitors (NRTIs) used for the treatment of HIV can a cause distal symmetrical peripheral polyneuropathy by disruption of mitochondrial metabolism. Treatment with acetyl-L-carnitine (ALCAR) has shown short-term symptomatic and histological improvement. Long-term effects have not been investigated. Purpose: To assess the subjective and objective degree of antiretroviral toxic neuropathy (ATN) during treatment with ALCAR. Method: A cohort of 21 patients with ATN who commenced treatment with ALCAR between March 1999 and October 2001 was reviewed after a mean of 4.3 years using standardized questionnaires and neurological examination. Results: Of the 21 patients, 2 had died and 3 were lost to follow-up. 16 patients were assessed. 10 were still on potentially neurotoxic drugs. 13 were still taking ALCAR. 9 were pain free. The most common symptom was numbness (mild, moderate, and severe in 12, 3, and 0 patients, respectively), followed by paraesthesia (8, 2, 2), pain (4, 3, 0), and burning (5, 2, 0). There was mildly reduced sensation in the toes of 8 patients. 13 patients reported that ALCAR had improved their symptoms very much or moderately, 2 reported no change, and 1 reported a moderate worsening. Conclusion: ALCAR led to long-term symptomatic improvement in most patients without the need to discontinue neurotoxic drugs. Although in this study there was no control group, this agent appears to be an effective pathogenesis-based treatment for ATN.

Selective vulnerability of the CA1 region of hippocampus to the indirect excitotoxic effects of malonic acid

The CA1 region of hippocampus is selectively vulnerable to a variety of insults, including hypoxia-ischemia and Alzheimer's disease, but the basis of this regional susceptibility is poorly understood. We examined the regional hippocampal sensitivity to mitochondrial metabolic disruption induced by malonate, an inhibitor of succinate dehydrogenase. The CA1 region was exquisitely sensitive to malonate and the dentate gyrus was extremely resistant; the CA3 region had intermediate sensitivity. This pattern of vulnerability is reminiscent of hypoxic-ischemic damage. Malonate damage was blocked by the N-methyl- d-aspartic acid (NMDA) antagonist, MK-801, but regional susceptibility to malonate did not correlate with the density of NMDA receptors. Instead, malonate toxicity was inversely correlated with activity of succinate dehydrogenase. Our results suggest that regional metabolic capacity may help to determine sensitivity to metabolic/excitotoxic insults such as hypoxia-ischemia.

The effect of malate on propionate mitochondrial toxicity

Propionic acidemia occasionally produces a toxic encephalopathy resembling Reye's syndrome, indicating disruption of mitochondrial metabolism. Liver mitochondria respiratory control ratios were reduced 46% by 5 mM propionate; inhibition correlated with matrix propionyl-CoA levels. L-Malate prevented the toxic effect of propionate and reduced the propionyl-CoA matrix concentration by 62%. The beneficial effect of L-malate is apparently due to stimulation of succinate efflux because the effect is blocked by benzylmalonate, an inhibitor of the dicarboxylate carrier. Matrix concentration of label from [1-14C]propionate was not affected by L-malate and/or benzylmalonate. L-Malate may be useful in the treatment of patients with propionic acidemia.

Propionate mitochondrial toxicity in liver and skeletal muscle: Acyl CoA levels

Propionic acidemia occasionally produces a toxic encephalopathy resembling Reye syndrome, indicating disruption of mitochondrial metabolism. Understanding the mitochondrial effect of propionate might clarify the pathophysiology. Liver mitochondria are inhibited by propionate (5 mM) while muscle mitochondria are not. Preincubation is required to inhibit liver mitochondria, suggesting that propionate is metabolized to propionyl CoA. Liver and skeletal muscle mitochondria incubated with [1-14C]propionate contain similar quantities of matrix isotope and release comparable [14C]CO2. However, only liver mitochondria accumulated significant propionyl CoA, which was largely (68%) synthesized from propionate. Carnitine reduced the level of liver matrix propionyl CoA. Inhibition of respiratory control ratios by propionate correlated with propionyl CoA levels. These results support the hypothesis that acyl CoA esters are toxic and that carnitine exerts its protective effect by converting acyl CoA esters to acylcarnitine esters.