Induction Of Mitochondrial Permeability Transition Research Articles

Cyclosporine-insensitive mode of cell death after prolonged myocardial ischemia: Evidence for sarcolemmal permeabilization as the pivotal step.

A prominent theory of cell death in myocardial ischemia/reperfusion (I/R) posits that the primary and pivotal step of irreversible cell injury is the opening of the mitochondrial permeability transition (MPT) pore. However, the predominantly positive evidence of protection against infarct afforded by the MPT inhibitor, Cyclosporine A (CsA), in experimental studies is in stark contrast with the overall lack of benefit found in clinical trials of CsA. One reason for the discrepancy might be the fact that relatively short experimental ischemic episodes (<1 hour) do not represent clinically-realistic durations, usually exceeding one hour. Here we tested the hypothesis that MPT is not the primary event of cell death after prolonged (60–80 min) episodes of global ischemia. We used confocal microcopy in Langendorff-perfused rabbit hearts treated with the electromechanical uncoupler, 2,3-Butanedione monoxime (BDM, 20 mM) to allow tracking of MPT and sarcolemmal permeabilization (SP) in individual ventricular myocytes. The time of the steepest drop in fluorescence of mitochondrial membrane potential (ΔΨm)-sensitive dye, TMRM, was used as the time of MPT (TMPT). The time of 20% uptake of the normally cell-impermeable dye, YO-PRO1, was used as the time of SP (TSP). We found that during reperfusion MPT and SP were tightly coupled, with MPT trending slightly ahead of SP (TSP-TMPT = 0.76±1.31 min; p = 0.07). These coupled MPT/SP events occurred in discrete myocytes without crossing cell boundaries. CsA (0.2 μM) did not reduce the infarct size, but separated SP and MPT events, such that detectable SP was significantly ahead of MPT (TSP -TMPT = -1.75±1.28 min, p = 0.006). Mild permeabilization of cells with digitonin (2.5–20 μM) caused coupled MPT/SP events which occurred in discrete myocytes similar to those observed in Control and CsA groups. In contrast, deliberate induction of MPT by titration with H2O2 (200–800 μM), caused propagating waves of MPT which crossed cell boundaries and were uncoupled from SP. Taken together, these findings suggest that after prolonged episodes of ischemia, SP is the primary step in myocyte death, of which MPT is an immediate and unavoidable consequence.

The antioxidant, aged garlic extract, exerts cytotoxic effects on wild-type and multidrug-resistant human cancer cells by altering mitochondrial permeability

Aged garlic extract (AGE) has been shown to possess therapeutic properties in cancer; however its mechanisms of action are unclear. In this study, we demonstrate by MTT assay that AGE exerts an anti-proliferative effect on a panel of both sensitive and multidrug-resistant (MDR) human cancer cell lines and enhances the effects of hyperthermia (42˚C) on M14 melanoma cells. The evaluation of the mitochondrial activity in whole cancer cells treated with AGE, performed by cytofluorimetric analysis in the presence of the lipophilic cationic fluorochrome JC-1, revealed the occurrence of dose-dependent mitochondrial membrane depolarization. Membrane potential was measured by the TPP+ selective electrode. In order to shed light on its mechanisms of action, the effects of AGE on isolated rat liver mitochondria were also examined. In this regard, AGE induced a mitochondrial membrane hyperpolarization of approximately 15mV through a mechanism that was similar to that observed with the ionophores, nigericin or salinomycin, by activating an exchange between endogenous K+ with exogenous H+. The prolonged incubation of the mitochondria with AGE induced depolarization and matrix swelling, indicative of mitochondrial permeability transition induction that, however, occurs through a different mechanism from the well-known one. In particular, the transition pore opening induced by AGE was due to the rearrangement of the mitochondrial membranes following the increased activity of the K+/H+ exchanger. On the whole, the findings of this study indicate that AGE exerts cytotoxic effects on cancer cells by altering mitochondrial permeability. In particular, AGE in the mitochondria activates K+/H+ exchanger, causes oxidative stress and induces mitochondrial permeability transition(MPT).

The dual role of mitochondrial superoxide in arsenite toxicity: Signaling at the boundary between apoptotic commitment and cytoprotection

Arsenite toxicity is in numerous cellular systems dependent on the formation of reactive oxygen and or nitrogen species. This is also true in U937 cells in which the metalloid selectively promotes the formation of mitochondrial superoxide (mitoO2−) rapidly converted to diffusible H2O2. We tested the hypothesis that, under the same conditions, mitoO2− also mediates the triggering of a parallel survival signaling. We found that a low concentration of the metalloid causes an early activation of nuclear factor erythroid 2 p45-related factor 2 (Nrf2), and a downstream signaling leading to enhanced GSH biosynthesis, via a mechanism sensitive to various treatments/strategies selectively preventing mitoO2− formation. Under the same conditions, the toxic effects mediated by arsenite, leading to delayed mitochondrial permeability transition (MPT)-dependent apoptosis, were also prevented. Additional studies revealed remarkable similarities in the kinetics of mitoO2− formation, MPT induction, Nrf2 activation and GSH biosynthesis, prior to the onset of apoptosis in a small portion of the cells. Importantly, mitoO2− formation, as well as the ensuing toxic events, were significantly potentiated and anticipated under conditions associated with inhibition of de novo GSH biosynthesis triggered by the metalloid through Nrf2 activation.We conclude that, in the arsenite toxicity paradigm under investigation, mitoO2− represents the only trigger of two opposite pathways leading to activation of the Nrf2 signaling and/or to a MPT-dependent apoptotic death. The first pathway, through enhanced GSH biosynthesis, mitigates the extent of further mitoO2− formation, thereby limiting and delaying an otherwise rapid and massive apoptotic death.

Mitochondrial toxicity induced by a thiourea gold(i) complex: mitochondrial permeability transition and respiratory deficit.

Gold(i) complexes have been widely used as antibacterial and antitumor agents because of their excellent biological activities. However, there are few reports on the study of gold(i) complexes at the subcellular level. Herein, we investigated the toxicity of a gold(i) complex (N,N'-disubstituted cyclic thiourea ligand) - AuTuCl - to isolated mitochondria via various methods. The results showed that AuTuCl induced mitochondrial swelling, elevated ROS generation and triggered collapse of the membrane potential, which indicated the induction of mitochondrial permeability transition (MPT). It also enhanced the permeability of H+ and K+ of the inner membrane and declined membrane fluidity, which might be the result of MPT. Moreover, AuTuCl impaired the mitochondrial respiratory chain and suppressed the activities of complexes II and IV in the respiratory chain. It also triggered the deficiency of ATP and the effusion of Cyt c, which were strictly related to respiration and apoptosis. These results indicated that AuTuCl severely affected the structure and function of mitochondria. It was proposed that MPT and impairment of the respiratory chain were responsible for the mitotoxicity of AuTuCl, thus causing energy deficiency and even apoptosis. This conceivable mechanism can serve as a clue for better understanding of the toxicology of AuTuCl.

Mitochondria are an Important Target in the Search for new Drugs for the Treatment of Alzheimer′s Disease and Senile Dementia

The review and summarizes own and literature data about the role of mitochondria as the important target in the search for drugs for the treatment of neurodegenerative diseases. Aging is a major risk factor for sporadic forms of various neurodegenerative diseases, including Alzheimer′s disease. One of the most argued and currently accepted theories is the Mitochondrial Free Radical Theory of Aging. Mitochondrial hypotheses of the development of sporadic forms of neurodegenerative diseases particularly Alzheimer′s disease, are closely connected with it. Impairments of mitochondrial functions lead to a decrease in their ability to regulate calcium homeostasis in the cell and to a decrease in the threshold for the induction of mitochondrial permeability transition (MPT) pores. MPT inhibitors can be considered as a promising approach to the treatment of neurodegenerative diseases, since these drugs can not only exhibit the properties of neuroprotectors, but also can provide normalization of synaptic activity due to increased calcium capacity of mitochondria. The review presents data on the number of MPT inhibitors, including endogenous compounds melatonin and N-acetylserotonin, their bioisosteric analogue Dimebon and a number of other compounds. The use of mitochondria as a basis for the formation of screening strategy for the search for compounds for the treatment of neurodegenerative diseases is of particular interest – both as a test of their potential toxicity, and as a basis for the creation of metabolic stimulants and drugs with neuroprotective and cognitive-stimulating effect.

Arsenite induces DNA damage via mitochondrial ROS and induction of mitochondrial permeability transition.

Arsenite is an established DNA-damaging agent and human carcinogen. We initially selected conditions in which the metalloid causes DNA strand scission in the absence of detectable apoptotic DNA degradation in U937 cells. This response was suppressed by catalase and by treatments (rotenone and ascorbic acid), or manipulations (respiration-deficient phenotype), preventing the mitochondrial formation of O2-. ( mitoO2-.). MitoO2-., and its dismutation product, H2 O2 , are therefore critically involved in the arsenite-dependent DNA-damaging response. We then established a link between mitoO2-./H2 O2 and mitochondrial permeability transition (MPT), and found that this second event also promoted the formation of DNA-damaging species. As a consequence, the DNA damage induced by arsenite, in addition to being abolished by the aforementioned treatments/manipulations, was also significantly reduced by the MPT inhibitor cyclosporin A (CsA). A CsA-sensitive induction of p53 mRNA expression was also detected. Finally, evidence of CsA-sensitive DNA strand scission was also obtained in MCF-7, HT22, and NCTC-2544 cells. MitoO2-./H2 O2 therefore directly mediates DNA damage induced by arsenite and indirectly promotes the formation of additional DNA-damaging species via the induction of MPT. © 2017 BioFactors, 43(5):673-684, 2017.

Mitochondrial permeability transition involves dissociation of F1FO ATP synthase dimers and C-ring conformation.

The impact of the mitochondrial permeability transition (MPT) on cellular physiology is well characterized. In contrast, the composition and mode of action of the permeability transition pore complex (PTPC), the supramolecular entity that initiates MPT, remain to be elucidated. Specifically, the precise contribution of the mitochondrial F1FO ATP synthase (or subunits thereof) to MPT is a matter of debate. We demonstrate that F1FO ATP synthase dimers dissociate as the PTPC opens upon MPT induction. Stabilizing F1FO ATP synthase dimers by genetic approaches inhibits PTPC opening and MPT Specific mutations in the F1FO ATP synthase c subunit that alter C-ring conformation sensitize cells to MPT induction, which can be reverted by stabilizing F1FO ATP synthase dimers. Destabilizing F1FO ATP synthase dimers fails to trigger PTPC opening in the presence of mutants of the c subunit that inhibit MPT The current study does not provide direct evidence that the C-ring is the long-sought pore-forming subunit of the PTPC, but reveals that PTPC opening requires the dissociation of F1FO ATP synthase dimers and involves the C-ring.

5 - Manganese Enhances Mitochondrial H2O2 Emission by Different Mechanisms

The mitochondrial respiratory chain is a major source of cellular reactive oxygen species (ROS). Recent data indicate that beside the established H 2 O 2 /O 2 ●– -generators NADH-ubiquinone- (complex I) and the ubiquinone-cytrochrome c -oxidoreductase (complex III), additional sources significantly contribute to the overall ROS production. These include the oxoglutarate dehydrogenase complex and the succinate dehydrogenase (complex II; SDH). The latter was shown to produce mainly H 2 O 2 from its FAD site. More recently manganese toxicity in the central nervous system has been linked to an elevated ROS production by complex II. However, the molecular mechanism as well as the distinct site of origin remained elusive. In this study with submitochondrial particles (SMP) and isolated rat heart mitochondria (RHM) we confirmed an inducing effect of manganese on ROS production by complex II. Furthermore, we tracked down H 2 O 2 /O 2 ●– -to be derived mainly from the ubiquinone binding site. In RHM, manganese additionally acted as a potent inducer of mitochondrial permeability transition, and hence potentiated the emission of H 2 O 2 from ‘substrate’ oxidoreductases of the Krebs cycle. Moreover, manganese generally led to an enhanced emission of hydrogen peroxide due to its accelerating effect on superoxide dismutation. These findings highlight the complex effects of manganese on mitochondrial H 2 O 2 emission. They also warrant further explorations aiming to understand/predict the functional consequences of H 2 O 2 /O 2 ●– formed at distinct sites of SDH.

2-Methylcitric acid impairs glutamate metabolism and induces permeability transition in brain mitochondria.

Accumulation of 2-methylcitric acid (2MCA) is observed in methylmalonic and propionic acidemias, which are clinically characterized by severe neurological symptoms. The exact pathogenetic mechanisms of brain abnormalities in these diseases are poorly established and very little has been reported on the role of 2MCA. In the present work we found that 2MCA markedly inhibited ADP-stimulated and uncoupled respiration in mitochondria supported by glutamate, with a less significant inhibition in pyruvate plus malate respiring mitochondria. However, no alterations occurred when α-ketoglutarate or succinate was used as respiratory substrates, suggesting a defect on glutamate oxidative metabolism. It was also observed that 2MCA decreased ATP formation in glutamate plus malate or pyruvate plus malate-supported mitochondria. Furthermore, 2MCA inhibited glutamate dehydrogenase activity at concentrations as low as 0.5 mM. Kinetic studies revealed that this inhibitory effect was competitive in relation to glutamate. In contrast, assays of osmotic swelling in non-respiring mitochondria suggested that 2MCA did not significantly impair mitochondrial glutamate transport. Finally, 2MCA provoked a significant decrease in mitochondrial membrane potential and induced swelling in Ca(2+)-loaded mitochondria supported by different substrates. These effects were totally prevented by cyclosporine A plus ADP or ruthenium red, indicating induction of mitochondrial permeability transition. Taken together, our data strongly indicate that 2MCA behaves as a potent inhibitor of glutamate oxidation by inhibiting glutamate dehydrogenase activity and as a permeability transition inducer, disturbing mitochondrial energy homeostasis. We presume that 2MCA-induced mitochondrial deleterious effects may contribute to the pathogenesis of brain damage in patients affected by methylmalonic and propionic acidemias. We propose that brain glutamate oxidation is disturbed by 2-methylcitric acid (2MCA), which accumulates in tissues from patients with propionic and methylmalonic acidemias because of a competitive inhibition of glutamate dehydrogenase (GDH) activity. 2MCA also induced mitochondrial permeability transition (PT) and decreased ATP generation in brain mitochondria. We believe that these pathomechanisms may be involved in the neurological dysfunction of these diseases.

Neuroprotective Effects of the Securinine-Analogues: Identification of Allomargaritarine as a Lead Compound.

Oxidative stress and mitochondrial disturbances are the common and important causative factors of aging, and play an important role in the late onset of sporadic neurodegenerative diseases, including Alzheimer disease (AD). Furthermore, emerging evidence from in vitro and in vivo disease models suggests that oxidative stress and increased vulnerability to induction of mitochondrial permeability transition leads to the pathogenesis of the neurological disorders. Towards the goals of developing effective neuroprotectors, this article describes the synthesis and neuroprotective studies of various derivatives of the naturally occurring alkaloid securinine, based on which a lead compound, allomargaritarine (a diastereomer of margaritarine), was identified as an effective therapeutic for neuroprotection. Allomargaritarine exhibits high antioxidant activity, and has significant mitoprotective effect on cellular models of neurodegeneration.

Sildenafil Ameliorates Advanced Glycation End Products-Induced Mitochondrial Dysfunction in HT-22 Hippocampal Neuronal Cells.

ObjectiveAccumulation of advanced glycation end-products (AGE) and mitochondrial glycation is importantly implicated in the pathological changes of the brain associated with diabetic complications, Alzheimer disease, and aging. The present study was undertaken to determine whether sildenafil, a type 5 phosphodiesterase type (PDE-5) inhibitor, has beneficial effect on neuronal cells challenged with AGE-induced oxidative stress to preserve their mitochondrial functional integrity.MethodsHT-22 hippocampal neuronal cells were exposed to AGE and changes in the mitochondrial functional parameters were determined. Pretreatment of cells with sildenafil effectively ameliorated these AGE-induced deterioration of mitochondrial functional integrity.ResultsAGE-treated cells lost their mitochondrial functional integrity which was estimated by their MTT reduction ability and intracellular ATP concentration. These cells exhibited stimulated generation of reactive oxygen species (ROS), disruption of mitochondrial membrane potential, induction of mitochondrial permeability transition, and release of the cytochrome C, activation of the caspase-3 accompanied by apoptosis. Western blot analyses and qRT-PCR demonstrated that sildenafil increased the expression level of the heme oxygenase-1 (HO-1). CoPP and bilirubin, an inducer of HO-1 and a metabolic product of HO-1, respectively, provided a similar protective effects. On the contrary, the HO-1 inhibitor ZnPP IX blocked the effect of sildenafil. Transfection with HO-1 siRNA significantly reduced the protective effect of sildenafil on the loss of MTT reduction ability and MPT induction in AGE-treated cells.ConclusionTaken together, our results suggested that sildenafil provides beneficial effect to protect the HT-22 hippocampal neuronal cells against AGE-induced deterioration of mitochondrial integrity, and upregulation of HO-1 is involved in the underlying mechanism.

Mitochondrial Respiration Is Associated with Alloxan-Induced Mitochondrial Permeability Transition

We previously showed that increased mitochondrial inner membrane permeability which is known as mitochondrial permeability transition (MPT) is triggered by adding succinate in the presence of the diabetogenic agent alloxan. Here, our aim was to investigate whether mitochondrial respiration is associated with alloxan-induced MPT. After mitochondria isolated from rat liver were incubated with alloxan at 37°C for 5 min, the addition of succinate immediately triggered the MPT in the presence of rotenone. However, little or no induction occurred at incubation temperatures below 25°C. Malate/glutamate also triggered MPT by alloxan in the absence of rotenone. In mitochondrial suspensions containing alloxan, succinate accelerated oxygen consumption that was completely inhibited by cyanide. These results suggest that mitochondrial respiration is associated with the alloxan-induced MPT. Alloxan radical production was investigated using ESR spectroscopy. Mitochondria incubated with succinate and alloxan elicited low signal intensity (radical formation) that increased significantly in the presence of cyanide. When the incubation of alloxan with mitochondria after the addition of succinate, a little intensity of the signal was observed, but it was remarkably increased after the addition of cyanide. Ubiquinone analogues inhibited the MPT induction. These results suggest that the initiation of MPT is associated with alloxan redox cycling via an electron transfer process at a quinone-binding site in respiratory mitochondria.

Uncoupling, metabolic inhibition and induction of mitochondrial permeability transition in rat liver mitochondria caused by the major long-chain hydroxyl monocarboxylic fatty acids accumulating in LCHAD deficiency

Patients with long-chain 3-hydroxy-acyl-CoA dehydrogenase (LCHAD) deficiency commonly present liver dysfunction whose pathogenesis is unknown. We studied the effects of long-chain 3-hydroxylated fatty acids (LCHFA) that accumulate in LCHAD deficiency on liver bioenergetics using mitochondrial preparations from young rats. We provide strong evidence that 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, the monocarboxylic acids that are found at the highest tissue concentrations in this disorder, act as metabolic inhibitors and uncouplers of oxidative phosphorylation. These conclusions are based on the findings that these fatty acids decreased ADP-stimulated (state 3) and uncoupled respiration, mitochondrial membrane potential and NAD(P)H content, and, in contrast, increased resting (state 4) respiration. We also verified that 3HTA and 3HPA markedly reduced Ca2+ retention capacity and induced swelling in Ca2+-loaded mitochondria. These effects were mediated by mitochondrial permeability transition (MPT) induction since they were totally prevented by the classical MPT inhibitors cyclosporin A and ADP, as well as by ruthenium red, a Ca2+ uptake blocker. Taken together, our data demonstrate that the major monocarboxylic LCHFA accumulating in LCHAD deficiency disrupt energy mitochondrial homeostasis in the liver. It is proposed that this pathomechanism may explain at least in part the hepatic alterations characteristic of the affected patients.

The calcium-ROS-pH triangle and mitochondrial permeability transition: challenges to mimic cardiac ischemia-reperfusion.

The calcium-ROS-pH triangle and mitochondrial permeability transition: challenges to mimic cardiac ischemia-reperfusion.

Donepezil attenuates Aβ-associated mitochondrial dysfunction and reduces mitochondrial Aβ accumulation in vivo and in vitro

The main purpose of the present study is to investigate the influence of donepezil, a well-known acetylcholinesterase (AChE) inhibitor, on amyloid-β (Aβ)-associated mitochondrial dysfunction, in order to gain a better understanding of the neuroprotective effects of this clinically used anti-Alzheimer's disease (AD) drug. First, our study verifies the ameliorative effects of donepezil on behavioral deficits in both working memory and anxiety in APP/PS1 double transgenic mice, at a time point that AChE is not inhibited. Meanwhile, we demonstrate that donepezil enhances the resistance of brain mitochondria of APP/PS1 mice to the induction of mitochondrial permeability transition (MPT) by calcium ions. Moreover, the level of mitochondrial Aβ in the brain of donepezil-treated APP/PS1 transgenic mice is significantly lower than that of vehicle-treated APP/PS1 mice. Our in vitro study using isolated mitochondria from rat brains, which is expected as an AChE-free subcellular system, further confirms the ameliorative effects of donepezil on oligomeric Aβ1-42 induced mitochondrial swelling and ATP reduction. In addition, donepezil treatment also significantly blocks the Aβ accumulation in the isolated mitochondria. Our study reported for the first time that the protective effects of donepezil against Aβ-associated mitochondrial dysfunction are closely associated with the reduction of Aβ accumulation in the mitochondria. Above observation led us to assume that, besides potent AChE inhibitory effect, other non-cholinergic mechanisms may be involved in the neuroprotective profiles of donepezil.

Targeting Mitochondria and Reactive Oxygen Species-Driven Pathogenesis in Diabetic Nephropathy

Diabetic kidney disease is one of the major microvascular complications of both type 1 and type 2 diabetes mellitus. Approximately 30% of patients with diabetes experience renal complications. Current clinical therapies can only mitigate the symptoms and delay the progression to end-stage renal disease, but not prevent or reverse it. Oxidative stress is an important player in the pathogenesis of diabetic nephropathy. The activity of reactive oxygen and nitrogen species (ROS/NS), which are by-products of the diabetic milieu, has been found to correlate with pathological changes observed in the diabetic kidney. However, many clinical studies have failed to establish that antioxidant therapy is renoprotective. The discovery that increased ROS/NS activity is linked to mitochondrial dysfunction, endoplasmic reticulum stress, inflammation, cellular senescence, and cell death calls for a refined approach to antioxidant therapy. It is becoming clear that mitochondria play a key role in the generation of ROS/NS and their consequences on the cellular pathways involved in apoptotic cell death in the diabetic kidney. Oxidative stress has also been associated with necrosis via induction of mitochondrial permeability transition. This review highlights the importance of mitochondria in regulating redox balance, modulating cellular responses to oxidative stress, and influencing cell death pathways in diabetic kidney disease. ROS/NS-mediated cellular dysfunction corresponds with progressive disease in the diabetic kidney, and consequently represents an important clinical target. Based on this consideration, this review also examines current therapeutic interventions to prevent ROS/NS-derived injury in the diabetic kidney. These interventions, mainly aimed at reducing or preventing mitochondrial-generated oxidative stress, improving mitochondrial antioxidant defense, and maintaining mitochondrial integrity, may deliver alternative approaches to halt or prevent diabetic kidney disease.

Some insight into the mode of cytotoxic action of organotin compounds with protective 2,6-di-tert-butylphenol fragments

A series of cytotoxic organotin compounds with Sn–S bonds of formulae Me2Sn(SR)2 (1); Et2Sn(SR)2 (2); (n-Bu)2Sn(SR)2 (3); Ph2Sn(SR)2 (4); R2Sn(SR)2 (5); Me3SnSR (6) Ph3SnSR (7) (R = 3,5-di-tert-butyl-4-hydroxyphenyl), synthesized recently, have been used to study the possible mechanisms of their cytotoxicity. With this aim we have investigated the influence of organotin compounds 1–7 as well as their precursor R2SnCl2 (8) on tubulin assembly in microtubules. Compounds 1, 6–8 reveal the statistically significant inhibition of tubulin polymerization without notable disturbances of microtubules structure. Docking simulations revealed the possibility of organotin compounds to bind in the paclitaxel or vinblastine sites on the tubulin surface, but the mechanism of action of the compounds could not be explained based solely on these data. The influence on mitochondrial potential and induction of mitochondrial permeability transition also have been studied. Organotin compound 7 depolarises the mitochondria and induces the mitochondrial permeability transition. These properties may be the main reason of its cytotoxicity. Since the organotins possess the protective 2,6-di-tert-butylphenol groups the antioxidant potential of these compounds as inhibitors of mitochondrial lipid peroxidation was studied. All compounds under investigation effectively inhibit the Fe3+-induced mitochondrial lipid peroxidation. The comparative study of structure-activity relationship in the lipid peroxidation inhibition was performed. The influence of the organic groups nature, the number of hindered phenol fragments in organotins on their toxic effect is discussed. The introduction of hindered phenol groups decreases the cytotoxicity of compounds. This result opens up the possibility of designing novel anticancer drugs that might possess lower undesirable toxicity against normal cells.

Regorafenib impairs mitochondrial functions, activates AMP-activated protein kinase, induces autophagy, and causes rat hepatocyte necrosis.

The tyrosine kinase inhibitor regorafenib was approved by regulatory agencies for cancer treatment, albeit with strong warnings of severe hepatotoxicity included in the product label. The basis of this toxicity is unknown; one possible mechanism, that of mitochondrial damage, was tested. In isolated rat liver mitochondria, regorafenib directly uncoupled oxidative phosphorylation (OXPHOS) and promoted calcium overload-induced swelling, which were respectively prevented by the recoupler 6-ketocholestanol (KC) and the mitochondrial permeability transition (MPT) pore blocker cyclosporine A (CsA). In primary hepatocytes, regorafenib uncoupled OXPHOS, disrupted mitochondrial inner membrane potential (MMP), and decreased cellular ATP at 1h, and triggered MPT at 3h, which was followed by necrosis but not apoptosis at 7h and 24h, all of which were abrogated by KC. The combination of the glycolysis enhancer fructose plus the mitochondrial ATPase synthase inhibitor oligomycin A abolished regorafenib induced necrosis at 7h. This effect was not seen at 24h nor with the fructose or oligomycin A separately. CsA in combination with trifluoperazine, both MPT blockers, showed similar effects. Two compensatory mechanisms, activation of AMP-activated protein kinase (AMPK) to ameliorate ATP shortage and induction of autophagy to remove dysfunctional mitochondria, were found to be mobilized. Hepatocyte necrosis was enhanced either by the AMPK inhibitor Compound C or the autophagy inhibitor chloroquine, while autophagy inducer rapamycin was strongly cytoprotective. Remarkably, all toxic effects were observed at clinically-relevant concentrations of 2.5-15μM. These data suggest that uncoupling of OXPHOS and the resulting ATP shortage and MPT induction are the key mechanisms for regorafenib induced hepatocyte injury, and AMPK activation and autophagy induction serve as pro-survival pathways against such toxicity.

Necroptosis: is there a role for mitochondria?

Once thought to be a random process of cell death, necrosis can proceed via a defined molecular mechanism and is integral to physiological and pathological states. In particular a form of necrosis called necroptosis has been the subject of intense investigation. Necroptosis is initiated by tumor necrosis factor-α (TNFα), which leads to the activation of the kinase receptor-interacting protein 1 (RIP1). RIP1 then binds with and activates RIP3 to form the necrosome. RIP3 in turn interacts with and activates the pseudokinase mixed lineage kinase domain-like (MLKL). This complex has then been proposed to induce necrotic death via the induction of mitochondrial dysfunction, with a variety of mechanisms being put forth including: production of mitochondrial reactive oxygen species (ROS), activation of the mitochondrial phosphatase PGAM5, or induction of mitochondrial permeability transition (MPT). However, recent evidence suggests that none of these are involved in necroptosis, and that mitochondria may in fact be dispensable for this process. Therefore, the purpose of this perspective is to discuss the current understanding of necroptosis, and more specifically, what role if any do mitochondria play in this mechanism of cell death.

Dimebon Attenuates the A&amp;#946;-Induced Mitochondrial Permeabilization

The currently available experimental data supports the hypothesis that the neuroprotective effect of dimebon is related to the protection of the brain-mitochondria from neurodegeneration. In this study, the influence of dimebon on mitochondria was investigated to gain a better understanding of the neuroprotective effects of this drug. Here, we demonstrate that dimebon enhances the resistance of the isolated rat brain and liver mitochondria to the induction of mitochondrial permeability transition (MPT) by calcium ions even in the presence of atractyloside, a MPT pore (MPTP) opener, but is ineffective against atractyloside-induced mitochondria swelling. Unlike cyclosporine A (CsA), a MPTP inhibitor, Dimebon does not influence the adenine nucleotide translocase (ANT) conformational changes and is not able to prevent the MPT of de-energized mitochondria. Using three different assays, and using amyloid-β peptide for inducing mitochondrial toxicity, we show that the influence of dimebon on the calcium retention capacity (CRC) of mitochondria depends on the mode of calcium addition. No obvious influence of dimebon on CRC was observed under the conditions of calcium infusion in the pump mode but the increase of CRC of rat brain mitochondria was observed when calcium was added in the bolus mode; the addition of calcium in the single pulse mode led to the increase of the lag period of calcium efflux from mitochondria. From these studies it is shown that dimebon is effective against amyloid-β (Aβ) potentiated mitochondrial swelling and decrease of calcium retention capacity (CRC) of the brain mitochondria.