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Sample records for mitochondria-targeted triphenylphosphonium-conjugated nitroxide

  1. A mitochondria-targeted nitroxide is reduced to its hydroxylamine by ubiquinol in mitochondria.

    PubMed

    Trnka, Jan; Blaikie, Frances H; Smith, Robin A J; Murphy, Michael P

    2008-04-01

    Piperidine nitroxides such as TEMPOL act as antioxidants in vivo due to their interconversion among nitroxide, hydroxylamine, and oxoammonium derivatives, but the mechanistic details of these reactions are unclear. As mitochondria are a significant site of piperidine nitroxide metabolism and action, we synthesized a mitochondria-targeted nitroxide, MitoTEMPOL, by conjugating TEMPOL to the lipophilic triphenylphosphonium cation. MitoTEMPOL was accumulated several hundred-fold into energized mitochondria where it was reduced to the hydroxylamine by direct reaction with ubiquinol. This reaction occurred by transfer of H() from ubiquinol to the nitroxide, with the ubisemiquinone radical product predominantly dismutating to ubiquinone and ubiquinol, together with a small amount reacting with oxygen to form superoxide. The piperidine nitroxides TEMPOL, TEMPO, and butylTEMPOL reacted similarly with ubiquinol in organic solvents but in mitochondrial membranes the rates varied in the order: MitoTEMPOL > butylTEMPOL > TEMPO > TEMPOL, which correlated with the extent of access of the nitroxide moiety to ubiquinol within the membrane. These findings suggest ways of using mitochondria-targeted compounds to modulate the coenzyme Q pool within mitochondria in vivo, and indicate that the antioxidant effects of mitochondria-targeted piperidine nitroxides can be ascribed to their corresponding hydroxylamines.

  2. A Mitochondria-Targeted Nitroxide/Hemigramicidin S Conjugate Protects Mouse Embryonic Cells Against Gamma Irradiation

    SciTech Connect

    Jiang Jianfei; Belikova, Natalia A.; Hoye, Adam T.; Zhao Qing; Epperly, Michael W.; Greenberger, Joel S.; Wipf, Peter; Kagan, Valerian E.

    2008-03-01

    Purpose: To evaluate the in vitro radioprotective effect of the mitochondria-targeted hemigramicidin S-conjugated 4-amino-2,2,6,6-tetramethyl-piperidine-N-oxyl (hemi-GS-TEMPO) 5-125 in {gamma}-irradiated mouse embryonic cells and adenovirus-12 SV40 hybrid virus transformed human bronchial epithelial cells BEAS-2B and explore the mechanisms involved in its radioprotective effect. Methods and Materials: Cells were incubated with 5-125 before (10 minutes) or after (1 hour) {gamma}-irradiation. Superoxide generation was determined by using dihydroethidium assay, and lipid oxidation was quantitated by using a fluorescence high-performance liquid chromatography-based Amplex Red assay. Apoptosis was characterized by evaluating the accumulation of cytochrome c in the cytosol and externalization of phosphatidylserine on the cell surface. Cell survival was measured by means of a clonogenic assay. Results: Treatment (before and after irradiation) of cells with 5-125 at low concentrations (5, 10, and 20 {mu}M) effectively suppressed {gamma}-irradiation-induced superoxide generation, cardiolipin oxidation, and delayed irradiation-induced apoptosis, evaluated by using cytochrome c release and phosphatidylserine externalization. Importantly, treatment with 5-125 increased the clonogenic survival rate of {gamma}-irradiated cells. In addition, 5-125 enhanced and prolonged {gamma}-irradiation-induced G{sub 2}/M phase arrest. Conclusions: Radioprotection/mitigation by hemi-GS-TEMPO likely is caused by its ability to act as an electron scavenger and prevent superoxide generation, attenuate cardiolipin oxidation in mitochondria, and hence prevent the release of proapoptotic factors from mitochondria. Other mechanisms, including cell-cycle arrest at the G{sub 2}/M phase, may contribute to the protection.

  3. A Topical Mitochondria-Targeted Redox-Cycling Nitroxide Mitigates Oxidative Stress-Induced Skin Damage.

    PubMed

    Brand, Rhonda M; Epperly, Michael W; Stottlemyer, J Mark; Skoda, Erin M; Gao, Xiang; Li, Song; Huq, Saiful; Wipf, Peter; Kagan, Valerian E; Greenberger, Joel S; Falo, Louis D

    2017-03-01

    Skin is the largest human organ, and it provides a first line of defense that includes physical, chemical, and immune mechanisms to combat environmental stress. Radiation is a prevalent environmental stressor. Radiation-induced skin damage ranges from photoaging and cutaneous carcinogenesis caused by UV exposure, to treatment-limiting radiation dermatitis associated with radiotherapy, to cutaneous radiation syndrome, a frequently fatal consequence of exposures from nuclear accidents. The major mechanism of skin injury common to these exposures is radiation-induced oxidative stress. Efforts to prevent or mitigate radiation damage have included development of antioxidants capable of reducing reactive oxygen species. Mitochondria are particularly susceptible to oxidative stress, and mitochondrial-dependent apoptosis plays a major role in radiation-induced tissue damage. We reasoned that targeting a redox cycling nitroxide to mitochondria could prevent reactive oxygen species accumulation, limiting downstream oxidative damage and preserving mitochondrial function. Here we show that in both mouse and human skin, topical application of a mitochondrially targeted antioxidant prevents and mitigates radiation-induced skin damage characterized by clinical dermatitis, loss of barrier function, inflammation, and fibrosis. Further, damage mitigation is associated with reduced apoptosis, preservation of the skin's antioxidant capacity, and reduction of irreversible DNA and protein oxidation associated with oxidative stress. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  4. Mitochondria-Targeted Nitroxide, Mito-CP, Suppresses Medullary Thyroid Carcinoma Cell Survival In Vitro and In Vivo

    PubMed Central

    Starenki, Dmytro

    2013-01-01

    Context: Medullary thyroid carcinoma (MTC) is a neuroendocrine tumor mainly caused by mutations in the RET proto-oncogene. For MTC therapy, the U.S. Food and Drug Administration recently approved vandetanib and cabozantinib, multikinase inhibitors targeting RET and other tyrosine kinase receptors of vascular endothelial growth factor, epidermal growth factor, or hepatocyte growth factor. Nevertheless, not all patients with the progressive MTC respond to these drugs, requiring the development of additional therapeutic modalities that have distinct activity. Objective: We aimed to evaluate mitochondria-targeted carboxy-proxyl (Mito-CP), a mitochondria-targeted redox-sensitive agent, for its tumor-suppressive efficacy against MTC. Design: In vitro cultures of 2 human MTC cell lines, TT and MZ-CRC-1, and TT xenografts in mice were treated with Mito-CP in comparison with vandetanib. The effects on cell survival/death, RET expression, mitochondrial integrity, and oxidative stress were determined. Results: Contrary to vandetanib, Mito-CP induced RET downregulation and strong cytotoxic effects in both cell lines in vitro, including caspase-dependent apoptosis. These effects were accompanied by mitochondrial membrane depolarization, decreased oxygen consumption, and increased oxidative stress in cells. Intriguingly, Mito-CP–induced cell death, but not RET downregulation, was partially inhibited by the reactive oxygen species scavenger, N-acetyl-cysteine, indicating that Mito-CP mediates tumor-suppressive effects via redox-dependent as well as redox-independent mechanisms. Orally administered Mito-CP effectively suppressed TT xenografts in mice, with an efficacy comparable to vandetanib and relatively low toxicity to animals. Conclusion: Our results suggest that Mito-CP can effectively suppress MTC cell growth/survival via a mechanism distinct from vandetanib effects. Mitochondrial targeting may be a potential strategy for MTC therapy. PMID:23509102

  5. Mitochondria-targeted nitroxide, Mito-CP, suppresses medullary thyroid carcinoma cell survival in vitro and in vivo.

    PubMed

    Starenki, Dmytro; Park, Jong-In

    2013-04-01

    Medullary thyroid carcinoma (MTC) is a neuroendocrine tumor mainly caused by mutations in the RET proto-oncogene. For MTC therapy, the U.S. Food and Drug Administration recently approved vandetanib and cabozantinib, multikinase inhibitors targeting RET and other tyrosine kinase receptors of vascular endothelial growth factor, epidermal growth factor, or hepatocyte growth factor. Nevertheless, not all patients with the progressive MTC respond to these drugs, requiring the development of additional therapeutic modalities that have distinct activity. We aimed to evaluate mitochondria-targeted carboxy-proxyl (Mito-CP), a mitochondria-targeted redox-sensitive agent, for its tumor-suppressive efficacy against MTC. In vitro cultures of 2 human MTC cell lines, TT and MZ-CRC-1, and TT xenografts in mice were treated with Mito-CP in comparison with vandetanib. The effects on cell survival/death, RET expression, mitochondrial integrity, and oxidative stress were determined. Contrary to vandetanib, Mito-CP induced RET downregulation and strong cytotoxic effects in both cell lines in vitro, including caspase-dependent apoptosis. These effects were accompanied by mitochondrial membrane depolarization, decreased oxygen consumption, and increased oxidative stress in cells. Intriguingly, Mito-CP-induced cell death, but not RET downregulation, was partially inhibited by the reactive oxygen species scavenger, N-acetyl-cysteine, indicating that Mito-CP mediates tumor-suppressive effects via redox-dependent as well as redox-independent mechanisms. Orally administered Mito-CP effectively suppressed TT xenografts in mice, with an efficacy comparable to vandetanib and relatively low toxicity to animals. Our results suggest that Mito-CP can effectively suppress MTC cell growth/survival via a mechanism distinct from vandetanib effects. Mitochondrial targeting may be a potential strategy for MTC therapy.

  6. Ubiquinol and plastoquinol triphenylphosphonium conjugates can carry electrons through phospholipid membranes.

    PubMed

    Rokitskaya, Tatyana I; Murphy, Michael P; Skulachev, Vladimir P; Antonenko, Yuri N

    2016-10-01

    Many mitochondria-targeted antioxidants (MTAs) that comprise a quinol moiety covalently attached through an aliphatic carbon chain to the lipophilic triphenylphosphonium cation are widely used for evaluating the role of mitochondria in pathological processes involving oxidative stress. The potency of MTAs to carry electrons across biological membranes and thereby mediate transmembrane redox processes was unknown. To assess this, we measured the rate of ferricyanide reduction inside liposomes by external ascorbate. Here, we show that MTAs containing ubiquinone (MitoQ series) or plastoquinone (SkQ series) can carry electrons through lipid membranes, with the rate being inversely proportional to the length of the hydrocarbon linker group. Furthermore, this process was stimulated by the hydrophobic anion tetraphenylborate suggesting that permeation of the cationic MTA through the membrane was the rate-limiting step of the process. This conclusion was supported by the observation that the rate of MTA-induced electron transfer was insensitive to nigericin, in contrast to electron transfer mediated by neutral quinone derivatives. These findings indicate that MTAs can be utilized to transfer electrons across lipid membranes and this may be applicable to the study of the electron-transport chain in mitochondria and other natural membranes exhibiting redox processes. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Mitochondria-Targeting Ceria Nanoparticles as Antioxidants for Alzheimer's Disease.

    PubMed

    Kwon, Hyek Jin; Cha, Moon-Yong; Kim, Dokyoon; Kim, Dong Kyu; Soh, Min; Shin, Kwangsoo; Hyeon, Taeghwan; Mook-Jung, Inhee

    2016-02-23

    Mitochondrial oxidative stress is a key pathologic factor in neurodegenerative diseases, including Alzheimer's disease. Abnormal generation of reactive oxygen species (ROS), resulting from mitochondrial dysfunction, can lead to neuronal cell death. Ceria (CeO2) nanoparticles are known to function as strong and recyclable ROS scavengers by shuttling between Ce(3+) and Ce(4+) oxidation states. Consequently, targeting ceria nanoparticles selectively to mitochondria might be a promising therapeutic approach for neurodegenerative diseases. Here, we report the design and synthesis of triphenylphosphonium-conjugated ceria nanoparticles that localize to mitochondria and suppress neuronal death in a 5XFAD transgenic Alzheimer's disease mouse model. The triphenylphosphonium-conjugated ceria nanoparticles mitigate reactive gliosis and morphological mitochondria damage observed in these mice. Altogether, our data indicate that the triphenylphosphonium-conjugated ceria nanoparticles are a potential therapeutic candidate for mitochondrial oxidative stress in Alzheimer's disease.

  8. Mitochondria targeting nano agents in cancer therapeutics

    PubMed Central

    Zhang, Xiao-Ying; Zhang, Pei-Ying

    2016-01-01

    Mitochondria have emerged as noteworthy therapeutic targets as their physiological functions are often altered in pathological conditions such as cancer. The electronic databases of MEDLINE, EMBASE and PubMed were searched for recent studies reporting the importance of mitochondria targeting nanoagents in cancer therapeutics. The concluding remarks of the above papers mostly confirmed the growing potential of these novel nanoagents in the area of anticancer research. Furthermore, numerous studies demonstrated the immense potential of nanocarriers in delivering mitochondria-acting compounds to their target site. Among the assemblage of nanomaterials, carbon nanotubes (CNTs) are becoming more prominent for drug delivery due to favorable attributes including their unique shape, which promotes cellular uptake, and large aspect ratio that facilitates conjugation of bioactive molecules on their surface. The present review focused on the current view of variable options available in mitochondria-targeting anticancer therapeutics. It may be concluded that improvements are essential for its establishment as a gold standard therapeutic option especially in the clinical setting. PMID:28105197

  9. The mitochondria-targeted imidazole substituted oleic acid 'TPP-IOA' affects mitochondrial bioenergetics and its protective efficacy in cells is influenced by cellular dependence on aerobic metabolism.

    PubMed

    Maddalena, Lucas A; Ghelfi, Mikel; Atkinson, Jeffrey; Stuart, Jeffrey A

    2017-01-01

    A variety of mitochondria-targeted small molecules have been invented to manipulate mitochondrial redox activities and improve function in certain disease states. 3-Hydroxypropyl-triphenylphosphonium-conjugated imidazole-substituted oleic acid (TPP-IOA) was developed as a specific inhibitor of cytochrome c peroxidase activity that inhibits apoptosis by preventing cardiolipin oxidation and cytochrome c release to the cytosol. Here we evaluate the effects of TPP-IOA on oxidative phosphorylation in isolated mitochondria and on mitochondrial function in live cells. We demonstrate that, at concentrations similar to those required to achieve inhibition of cytochrome c peroxidase activity, TPP-IOA perturbs oxidative phosphorylation in isolated mitochondria. In live SH-SY5Y cells, TPP-IOA partially collapsed mitochondrial membrane potential, caused extensive fragmentation of the mitochondrial network, and decreased apparent mitochondrial abundance within 3h of exposure. Many cultured cell lines rely primarily on aerobic glycolysis, potentially making them less sensitive to small molecules disrupting oxidative phosphorylation. We therefore determined the anti-apoptotic efficacy of TPP-IOA in SH-SY5Y cells growing in glucose or in galactose, the latter of which increases reliance on oxidative phosphorylation for ATP supply. The anti-apoptotic activity of TPP-IOA that was observed in glucose media was not seen in galactose media. It therefore appears that, at concentrations required to inhibit cytochrome c peroxidase activity, TPP-IOA perturbs oxidative phosphorylation. In light of these data it is predicted that potential future therapeutic applications of TPP-IOA will be restricted to highly glycolytic cell types with limited reliance on oxidative phosphorylation. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Neuroprotection and Anti-Epileptogenesis with Mitochondria-Targeted Antioxidant

    DTIC Science & Technology

    2016-06-01

    Award Number: W81XWH-12-1-0258 TITLE: Neuroprotection and Anti-Epileptogenesis with Mitochondria -Targeted Antioxidant PRINCIPAL INVESTIGATOR...SUBTITLE Neuroprotection and Anti-Epileptogenesis with Mitochondria -Targeted Antioxidant 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-12-1-0258 5c

  11. Antiproliferative effects of mitochondria-targeted cationic antioxidants and analogs: Role of mitochondrial bioenergetics and energy-sensing mechanism

    PubMed Central

    Cheng, Gang; Zielonka, Jacek; McAllister, Donna; Hardy, Micael; Ouari, Olivier; Joseph, Joy; Dwinell, Michael B.; Kalyanaraman, Balaraman

    2015-01-01

    One of the proposed mechanisms for tumor proliferation involves redox signaling mediated by reactive oxygen species such as superoxide and hydrogen peroxide generated at moderate levels. Thus, the antiproliferative and anti-tumor effects of certain antioxidants were attributed to their ability to mitigate intracellular reactive oxygen species (ROS). Recent reports support a role for mitochondrial ROS in stimulating tumor cell proliferation. In this study, we compared the antiproliferative effects and the effects on mitochondrial bioenergetic functions of a mitochondria-targeted cationic carboxyproxyl nitroxide (Mito-CP), exhibiting superoxide dismutase (SOD)-like activity and a synthetic cationic acetamide analog (Mito-CP-Ac) lacking the nitroxide moiety responsible for the SOD activity. Results indicate that both Mito-CP and Mito-CP-Ac potently inhibited tumor cell proliferation. Both compounds altered mitochondrial and glycolytic functions, and intracellular citrate levels. Both Mito-CP and Mito-CP-Ac synergized with 2-deoxy-glucose (2-DG) to deplete intracellular ATP, inhibit cell proliferation and induce apoptosis in pancreatic cancer cells. We conclude that mitochondria-targeted cationic agents inhibit tumor proliferation via modification of mitochondrial bioenergetics pathways rather than by dismutating and detoxifying mitochondrial superoxide. PMID:26004344

  12. Antiproliferative effects of mitochondria-targeted cationic antioxidants and analogs: Role of mitochondrial bioenergetics and energy-sensing mechanism.

    PubMed

    Cheng, Gang; Zielonka, Jacek; McAllister, Donna; Hardy, Micael; Ouari, Olivier; Joseph, Joy; Dwinell, Michael B; Kalyanaraman, Balaraman

    2015-08-28

    One of the proposed mechanisms for tumor proliferation involves redox signaling mediated by reactive oxygen species such as superoxide and hydrogen peroxide generated at moderate levels. Thus, the antiproliferative and anti-tumor effects of certain antioxidants were attributed to their ability to mitigate intracellular reactive oxygen species (ROS). Recent reports support a role for mitochondrial ROS in stimulating tumor cell proliferation. In this study, we compared the antiproliferative effects and the effects on mitochondrial bioenergetic functions of a mitochondria-targeted cationic carboxyproxyl nitroxide (Mito-CP), exhibiting superoxide dismutase (SOD)-like activity and a synthetic cationic acetamide analog (Mito-CP-Ac) lacking the nitroxide moiety responsible for the SOD activity. Results indicate that both Mito-CP and Mito-CP-Ac potently inhibited tumor cell proliferation. Both compounds altered mitochondrial and glycolytic functions, and intracellular citrate levels. Both Mito-CP and Mito-CP-Ac synergized with 2-deoxy-glucose (2-DG) to deplete intracellular ATP, inhibit cell proliferation and induce apoptosis in pancreatic cancer cells. We conclude that mitochondria-targeted cationic agents inhibit tumor proliferation via modification of mitochondrial bioenergetics pathways rather than by dismutating and detoxifying mitochondrial superoxide.

  13. Mitochondria-targeted nutraceuticals in sports medicine: a new perspective.

    PubMed

    Ostojic, Sergej M

    2017-01-01

    Since mitochondria have been recognized as the cells' key organelles involved in the energy utilization during exercise, targeting the organelle with specifically designed compounds (mitochondria-targeted nutraceuticals, MTNs) may have a great promise in the prevention and treatment of heavy exercise-related mitochondrial dysfunction. In vitro studies suggested that MTNs have antioxidant effects at the molecular level, and might boost mitochondrial biogenesis and organelle bioenergetics, with both processes are known to positively affect exercise performance and recovery. However, while there are a number of different MTNs evaluated for a potential benefit as a therapy for mitochondria-related diseases and conditions, only few human studies evaluated the possible impact of novel MTNs in the field of sports medicine. This mini review summarizes recent research findings regarding the efficacy of different mitochondria-targeted nutritional agents, emphasizing their roles in sports medicine.

  14. Mitochondria-Targeted Protective Compounds in Parkinson's and Alzheimer's Diseases

    PubMed Central

    Fernández-Moriano, Carlos; González-Burgos, Elena; Gómez-Serranillos, M. Pilar

    2015-01-01

    Mitochondria are cytoplasmic organelles that regulate both metabolic and apoptotic signaling pathways; their most highlighted functions include cellular energy generation in the form of adenosine triphosphate (ATP), regulation of cellular calcium homeostasis, balance between ROS production and detoxification, mediation of apoptosis cell death, and synthesis and metabolism of various key molecules. Consistent evidence suggests that mitochondrial failure is associated with early events in the pathogenesis of ageing-related neurodegenerative disorders including Parkinson's disease and Alzheimer's disease. Mitochondria-targeted protective compounds that prevent or minimize mitochondrial dysfunction constitute potential therapeutic strategies in the prevention and treatment of these central nervous system diseases. This paper provides an overview of the involvement of mitochondrial dysfunction in Parkinson's and Alzheimer's diseases, with particular attention to in vitro and in vivo studies on promising endogenous and exogenous mitochondria-targeted protective compounds. PMID:26064418

  15. A mitochondria-targeted derivative of ascorbate: MitoC

    PubMed Central

    Finichiu, Peter G.; Larsen, David S.; Evans, Cameron; Larsen, Lesley; Bright, Thomas P.; Robb, Ellen L.; Trnka, Jan; Prime, Tracy A.; James, Andrew M.; Smith, Robin A.J.; Murphy, Michael P.

    2015-01-01

    Mitochondrial oxidative damage contributes to a wide range of pathologies. One therapeutic strategy to treat these disorders is targeting antioxidants to mitochondria by conjugation to the lipophilic triphenylphosphonium (TPP) cation. To date only hydrophobic antioxidants have been targeted to mitochondria; however, extending this approach to hydrophilic antioxidants offers new therapeutic and research opportunities. Here we report the development and characterization of MitoC, a mitochondria-targeted version of the hydrophilic antioxidant ascorbate. We show that MitoC can be taken up by mitochondria, despite the polarity and acidity of ascorbate, by using a sufficiently hydrophobic link to the TPP moiety. MitoC reacts with a range of reactive species, and within mitochondria is rapidly recycled back to the active ascorbate moiety by the glutathione and thioredoxin systems. Because of this accumulation and recycling MitoC is an effective antioxidant against mitochondrial lipid peroxidation and also decreases aconitase inactivation by superoxide. These findings show that the incorporation of TPP function can be used to target polar and acidic compounds to mitochondria, opening up the delivery of a wide range of bioactive compounds. Furthermore, MitoC has therapeutic potential as a new mitochondria-targeted antioxidant, and is a useful tool to explore the role(s) of ascorbate within mitochondria. PMID:26453920

  16. Mitochondria-targeted antioxidant effects of S(-) and R(+) pramipexole

    PubMed Central

    2010-01-01

    Background Pramipexole exists as two isomers. The S(-) enantiomer is a potent D3/D2 receptor agonist and is extensively used in the management of PD. In contrast, the R(+) enantiomer is virtually devoid of any of the DA agonist effects. Very limited studies are available to characterize the pharmacological spectrum of the R(+) enantiomer of pramipexole. Results Using differentiated SH-SY5Y neuroblastoma cells as an experimental model, here we show that S(-) and R(+) pramipexole are endowed with equipotent efficacy in preventing cell death induced by H2O2 and inhibiting mitochondrial reactive oxygen species generation. Both pramipexole enantiomers prevented mitochondrial ROS generation with a potency about ten times higher then that elicited for neuroprotection. Conclusions These results support the concept of both S(-) and R(+) pramipexole enantiomers as mitochondria-targeted antioxidants and suggest that the antioxidant, neuroprotective activity of these drugs is independent of both the chiral 6-propylamino group in the pramipexole molecule and the DA receptor stimulation. PMID:20137065

  17. Mitochondria-targeted antioxidants as highly effective antibiotics.

    PubMed

    Nazarov, Pavel A; Osterman, Ilya A; Tokarchuk, Artem V; Karakozova, Marina V; Korshunova, Galina A; Lyamzaev, Konstantin G; Skulachev, Maxim V; Kotova, Elena A; Skulachev, Vladimir P; Antonenko, Yuri N

    2017-05-03

    Mitochondria-targeted antioxidants are known to alleviate mitochondrial oxidative damage that is associated with a variety of diseases. Here, we showed that SkQ1, a decyltriphenyl phosphonium cation conjugated to a quinone moiety, exhibited strong antibacterial activity towards Gram-positive Bacillus subtilis, Mycobacterium sp. and Staphylococcus aureus and Gram-negative Photobacterium phosphoreum and Rhodobacter sphaeroides in submicromolar and micromolar concentrations. SkQ1 exhibited less antibiotic activity towards Escherichia coli due to the presence of the highly effective multidrug resistance pump AcrAB-TolC. E. coli mutants lacking AcrAB-TolC showed similar SkQ1 sensitivity, as B. subtilis. Lowering of the bacterial membrane potential by SkQ1 might be involved in the mechanism of its bactericidal action. No significant cytotoxic effect on mammalian cells was observed at bacteriotoxic concentrations of SkQ1. Therefore, SkQ1 may be effective in protection of the infected mammals by killing invading bacteria.

  18. Design, Synthesis, and Some Aspects of the Biological Activity of Mitochondria-Targeted Antioxidants.

    PubMed

    Korshunova, G A; Shishkina, A V; Skulachev, M V

    2017-07-01

    This review summarizes for the first time data on the design and synthesis of biologically active compounds of a new generation - mitochondria-targeted antioxidants, which are natural (or synthetic) p-benzoquinones conjugated via a lipophilic linker with (triphenyl)phosphonium or ammonium cations with delocalized charge. It also describes the synthesis of mitochondria-targeted antioxidants - uncouplers of oxidative phosphorylation - based on fluorescent dyes.

  19. Mitochondria-targeted antioxidants for treatment of Parkinson's disease: preclinical and clinical outcomes.

    PubMed

    Jin, Huajun; Kanthasamy, Arthi; Ghosh, Anamitra; Anantharam, Vellareddy; Kalyanaraman, Balaraman; Kanthasamy, Anumantha G

    2014-08-01

    Parkinson's disease is a progressive neurodegenerative disease in the elderly, and no cure or disease-modifying therapies exist. Several lines of evidence suggest that mitochondrial dysfunction and oxidative stress have a central role in the dopaminergic neurodegeneration of Parkinson's disease. In this context, mitochondria-targeted therapies that improve mitochondrial function may have great promise in the prevention and treatment of Parkinson's disease. In this review, we discuss the recent developments in mitochondria-targeted antioxidants and their potential beneficial effects as a therapy for ameliorating mitochondrial dysfunction in Parkinson's disease.

  20. MITOCHONDRIA-TARGETED ANTIOXIDANTS FOR TREATMENT OF PARKINSON’S DISEASE: PRECLINICAL AND CLINICAL OUTCOMES

    PubMed Central

    Jin, Huajun; Kanthasamy, Arthi; Ghosh, Anamitra; Anantharam, Vellareddy; Kalyanaraman, Balaraman; Kanthasamy, Anumantha G.

    2013-01-01

    Parkinson’s disease (PD) is a progressive neurodegenerative disease in the elderly, and no cure or disease-modifying therapies exist. Several lines of evidence suggest that mitochondrial dysfunction and oxidative stress have a central role in the dopaminergic neurodegeneration of PD. In this context, mitochondria-targeted therapies that improve mitochondrial function may have great promise in the prevention and treatment of PD. In this review, we discuss the recent developments in mitochondria-targeted antioxidants and their potential beneficial effects as a therapy for ameliorating mitochondrial dysfunction in PD. PMID:24060637

  1. Mito-methyl coumarin, a novel mitochondria-targeted drug with great antitumor potential was synthesized.

    PubMed

    Wang, Huanan; Xu, Wenqing

    2017-07-15

    Due to higher transmembrane potential of tumor cells, enhanced accumulation of cationic drugs in tumor mitochondria has been attributed to a higher (more negative inside) mitochondrial transmembrane potential compared with normal cells, emerging researchers are focus on developing mitochondria-targeted antitumor drugs. Coumarins showed great potential on antitumor, but mitochondria-targeted coumarin derivatives have not been reported. In the present study, we synthesized mitochondria-targeted-methyl coumarin (mito-methyl coumarin) through coupling 6-methyl coumarin to TPP. We confirmed that mito-methyl coumarin inhibited HeLa cells proliferation selectively, induced ROS generation, reduced mitochondrial membrane potential, promoted mitochondria Ca(2+) accumulation, decreased mitochondria mass and induced HeLa cells apoptosis, but methyl coumarin did not. These results demonstrate that we succeed in synthesizing a novel mitochondria-targeted drug, mito-methyl coumarin, which is effective in inhibiting HeLa cells proliferation and inducing HeLa cells apoptosis through promoting ROS generation and mitochondria Ca(2+) accumulation. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Mitochondria-targeted antioxidant SkQ1 improves spermatogenesis in Immp2l mutant mice.

    PubMed

    Jiang, Yaodong; Liu, Chunlian; Lei, Bin; Xu, Xian; Lu, Baisong

    2017-07-13

    Previous studies have confirmed that spermatogenesis in homozygous Immp2l mutant male mice was normal at the age of 6 months, but was significantly abnormal at the age of 13 months. Meanwhile, oxidative stress is reported to be involved in spermatogenic impairment in old mutant mice. However, it is unclear whether antioxidant treatment is a suitable intervention for improving spermatogenesis in old mutant mice. This study sought to investigate the effect of mitochondria-targeted antioxidant SkQ1 on spermatogenesis in homozygous Immp2l mutant mice. Immp2l mutant mice were treated with the mitochondria-targeted antioxidant SkQ1 from the age of 6 weeks until 13 months. SkQ1 treatment significantly improved spermatogenesis in old Immp2 l mutant mice. Moreover, SkQ1 treatment improved the morphology of testicular seminiferous tubules, significantly reduced the apoptosis of germ cells and increased the level of GPX4 expression in old Immp2 l mutant mice. In conclusion, our data suggest that the mitochondria-targeted antioxidant SkQ1 is effective in improving spermatogenesis in Immp2 l mutant mice and might be used for the treatment of male infertility. © 2017 Blackwell Verlag GmbH.

  3. Mitochondria-Targeting Polydopamine Nanoparticles To Deliver Doxorubicin for Overcoming Drug Resistance.

    PubMed

    Li, Wen-Qing; Wang, Zhigang; Hao, Sijie; He, Hongzhang; Wan, Yuan; Zhu, Chuandong; Sun, Li-Ping; Cheng, Gong; Zheng, Si-Yang

    2017-05-24

    Mitochondria play a critical role in diverse cellular processes, such as energy production and apoptosis regulation. The mitochondria-targeted drug delivery is becoming a potential novel strategy for overcoming drug resistance in cancer therapy. Herein, we synthesize nature-inspired dopamine-derived polydopamine (PDA) nanoparticles. Using triphenylphosphonium (TPP) as the mitochondrial penetration molecule to improve the target efficiency, we synthesize poly(ethylene glycol) (PEG)-modified PDA (PDA-PEG) and TPP-functionalized PEG-modified PDA (PDA-PEG-TPP) nanoparticles. Then anticancer drug doxorubicin (DOX) was loaded on PDA-PEG and PDA-PEG-TPP (PDA-PEG-DOX and PDA-PEG-TPP-DOX) nanoparticles, which are apt to deliver DOX to cell nuclei and mitochondria, respectively. To mimic the repeated anticancer drug treatment in clinical cases, we repeatedly treated the MDA-MD-231 cancer cells for a long time using DOX-loaded nanoparticles and find that the mitochondria targeting PDA-PEG-TPP-DOX has higher potential to overcome the drug resistance than the regular delivery nanoparticles PDA-PEG-DOX. These results indicate the promising potential of applying PDA-PEG-TPP-DOX nanoparticles in mitochondria-targeted drug delivery to overcome the drug resistance in long-time anticancer chemotherapy.

  4. Radioprotective effects of mitochondria-targeted antioxidant SkQR1.

    PubMed

    Fetisova, Elena K; Antoschina, Margarita M; Cherepanynets, Varvara D; Izumov, Denis S; Kireev, Igor I; Kireev, Roman I; Lyamzaev, Konstantin G; Riabchenko, Nikolay I; Chernyak, Boris V; Skulachev, Vladimir P

    2015-01-01

    We show here that mitochondria-targeted antioxidant composed of plastoquinone conjugated through hydrocarbon linker with cationic rhodamine 19 (SkQR1) protected against nuclear DNA damage induced by gamma radiation in K562 erythroleukemia cells. We also demonstrate that SkQR1 prevented the early (1 h postirradiation) accumulation of phosphorylated histone H2AX (γ-H2AX) an indicator of DNA double-strand break formation, as well as the radiation-induced increase in chromosomal aberrations. These data suggested that nuclear DNA damage induced by gamma radiation may be mediated by mitochondrial reactive oxygen species (ROS) production. We show that SkQR1 suppressed delayed accumulation of ROS 32 h after irradiation probably by inhibiting mitochondrial ROS-induced ROS release mechanisms. This suggests that mitochondria-targeted antioxidants may protect cells from the late consequences of radiation exposure related to delayed oxidative stress. We have previously reported that SkQRl is the substrate of multidrug resistance pump P-glycoproten (Pgp 170) and selectively protects Pgp 170-negative cells against oxidative stress. In line with this finding, we demonstrate here that SkQR1 did not protect Pgp170-positive K562 subline against DNA damage induced by gamma radiation. The selective radioprotection of normal Pgp 170-negative cells by mitochondria-targeted antioxidants could be a promising strategy to increase the efficiency of radiotherapy for multidrug-resistant tumors.

  5. Protective Effects of Melatonin and Mitochondria-targeted Antioxidants Against Oxidative Stress: A Review.

    PubMed

    Ramis, M R; Esteban, S; Miralles, A; Tan, Dun-Xian; Reiter, R J

    2015-01-01

    Oxidative damage is related to aging and a wide range of human disorders. Mitochondria are in large part responsible for free radical production and they are also main targets of the attack of these toxic molecules. The resulting deleterious effects of the damage to mitochondria can be prevented by antioxidants. Melatonin is an endogenously-produced indoleamine that modulates numerous functions, including mitochondria-related functions; this result from its capacity to penetrate all morphophysiological barriers and to enter all subcellular compartments due to its amphiphilic nature. Furthermore, this indoleamine and its metabolites are powerful antioxidants and scavengers of free radicals, protecting cellular membranes, the electron transport chain and mitochondrial DNA from oxidative damage. These properties may make melatonin a potent protector against a variety of free radical-related diseases. By comparison, other conventional antioxidants have less efficacy due to their limited access to the mitochondria. In recent years, research has focused on the advancement of mitochondria-targeted antioxidants, such as MitoQ (composed by the lipophilic triphenylphosphonium cation conjugated to the endogenous antioxidant coenzyme Q10) and MitoE (composed by the triphenylphosphonium cation attached to the antioxidant α-tocopherol). Mitochondria-targeted antioxidants accumulate in several hundred-fold greater concentrations within mitochondria and protect these critical organelles from oxidative damage. Melatonin also seems to be a mitochondria-targeted antioxidant and has similar protective actions as the synthetic antioxidants. Further work is required to determine the therapeutic properties of these antioxidants in ameliorating diseases related to mitochondrial dysfunction.

  6. A fluorescent carbon-dots-based mitochondria-targetable nanoprobe for peroxynitrite sensing in living cells.

    PubMed

    Wu, Xiaoxue; Sun, Shan; Wang, Yuhui; Zhu, Jiali; Jiang, Kai; Leng, Yumin; Shu, Qinghai; Lin, Hengwei

    2017-04-15

    Mitochondria, the power generators in cell, are a primary organelle of oxygen consumption and a main source of reactive oxygen/nitrogen species (ROS/RNS). Peroxynitrite (ONOO(-)), known as a kind of RNS, has been considered to be a significant factor in many cell-related biological processes, and there is great desire to develop fluorescent probes that can sensitively and selectively detect peroxynitrite in living cells. Herein, we developed a fluorescent carbon-dots (C-dots) based mitochondria-targetable nanoprobe with high sensitivity and selectivity for peroxynitrite sensing in living cells. The C-dots with its surface rich in amino groups was synthesized using o-phenylenediamine as carbon precursor, and it could be covalently conjugated with a mitochondria-targeting moiety, i.e. triphenylphosphonium (TPP). In the presence of peroxynitrite, the fluorescence of the constructed nanoprobe (C-dots-TPP) was efficiently quenched via a mechanism of photoinduced electron transfer (PET). The nanoprobe exhibited relatively high sensitivity (limit of detection: 13.5nM) and selectivity towards peroxynitrite in aqueous buffer. The performance of the nanoprobe for fluorescence imaging of peroxynitrite in mitochondria was investigated. The results demonstrated that the nanoprobe showed fine mitochondria-targeting ability and imaging contrast towards peroxynitrite in living cells. We anticipate that the proposed nanoprobe will provide a facile tool to explore the role played by peroxynitrite in cytobiology. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Ruthenium(II) polypyridyl complexes as mitochondria-targeted two-photon photodynamic anticancer agents.

    PubMed

    Liu, Jiangping; Chen, Yu; Li, Guanying; Zhang, Pingyu; Jin, Chengzhi; Zeng, Leli; Ji, Liangnian; Chao, Hui

    2015-07-01

    Clinical acceptance of photodynamic therapy is currently hindered by poor depth efficacy and inefficient activation of the cell death machinery in cancer cells during treatment. To address these issues, photoactivation using two-photon absorption (TPA) is currently being examined. Mitochondria-targeted therapy represents a promising approach to target tumors selectively and may overcome the resistance in current anticancer therapies. Herein, four ruthenium(II) polypyridyl complexes (RuL1-RuL4) have been designed and developed to act as mitochondria-targeted two-photon photodynamic anticancer agents. These complexes exhibit very high singlet oxygen quantum yields in methanol (0.74-0.81), significant TPA cross sections (124-198 GM), remarkable mitochondrial accumulation, and deep penetration depth. Thus, RuL1-RuL4 were utilized as one-photon and two-photon absorbing photosensitizers in both monolayer cells and 3D multicellular spheroids (MCSs). These Ru(II) complexes were almost nontoxic towards cells and 3D MCSs in the dark and generate sufficient singlet oxygen under one- and two-photon irradiation to trigger cell death. Remarkably, RuL4 exhibited an IC50 value as low as 9.6 μM in one-photon PDT (λirr = 450 nm, 12 J cm(-2)) and 1.9 μM in two-photon PDT (λirr = 830 nm, 800 J cm(-2)) of 3D MCSs; moreover, RuL4 is an order of magnitude more toxic than cisplatin in the latter test system. The combination of mitochondria-targeting and two-photon activation provides a valuable paradigm to develop ruthenium(II) complexes for PDT applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Contradictory Effects of Mitochondria- and Non-mitochondria-targeted Antioxidants on Hepatocarcinogenesis by Altering DNA Repair.

    PubMed

    Wang, Bibo; Fu, Jing; Yu, Ting; Xu, An; Qin, Wenhao; Yang, Zhishi; Chen, Yao; Wang, Hongyang

    2017-09-12

    Conflicting effects of antioxidant supplementation on cancer prevention or promotion is of great concern to healthy people and cancer patients. Despite recent studies about antioxidants accelerating the progression of lung cancer and melanoma, it does not fully deny antioxidants for cancer prevention. Both tumor and antioxidants types influence the actual efficacy. However, little is known about the impact of different types of antioxidants on primary hepatocellular carcinoma (HCC), including non-mitochondrial- and mitochondrial-targeted antioxidants. Based on the mouse models of chemical hepatocarcinogenesis, we showed that administration of non-mitochondria-targeted antioxidants N-acetylcysteine (NAC) and the soluble vitamin E analog Trolox prevented tumorigenesis, whereas administration of mitochondria-targeted antioxidants SS-31 (the mitochondria targeted peptide) and Mito-Q (a derivative of ubiquinone) encouraged tumorigenesis. RNA sequencing revealed that NAC and SS-31 cause highly different changes in oxidation-reduction state and DNA damage response. Remarkably, in diethylnitrosamine (DEN)-treated primary hepatocytes, NAC and Trolox alleviated DNA damage by activating ATM/ATR for DNA repair while SS-31 and MitoQ aggravated damage by inactivating them. Interestingly, partial recovery of SS-31-scavengened mitochondrial ROS (mtROS) could alleviate SS-31-aggravated DNA damage. Localization of ATM between mitochondria and nuclei was changed after NAC and SS-31 treatment. Furthermore, blockage of p-ATR led to the recurrence of NAC-ameliorated DEN HCC. In contrast, reactivation of p-ATR blocked SS-31-promoted DEN HCC. These results demonstrate that the type of antioxidants plays a previously unappreciated role in hepatocarcinogenesis, and provide a mechanistic rationale for exploring the therapeutic use of antioxidants for liver cancer. This article is protected by copyright. All rights reserved. © 2017 by the American Association for the Study of Liver Diseases.

  9. Mitochondria-targeted vitamin E analogs inhibit breast cancer cell energy metabolism and promote cell death.

    PubMed

    Cheng, Gang; Zielonka, Jacek; McAllister, Donna M; Mackinnon, A Craig; Joseph, Joy; Dwinell, Michael B; Kalyanaraman, Balaraman

    2013-06-13

    Recent research has revealed that targeting mitochondrial bioenergetic metabolism is a promising chemotherapeutic strategy. Key to successful implementation of this chemotherapeutic strategy is the use of new and improved mitochondria-targeted cationic agents that selectively inhibit energy metabolism in breast cancer cells, while exerting little or no long-term cytotoxic effect in normal cells. In this study, we investigated the cytotoxicity and alterations in bioenergetic metabolism induced by mitochondria-targeted vitamin E analog (Mito-chromanol, Mito-ChM) and its acetylated ester analog (Mito-ChMAc). Assays of cell death, colony formation, mitochondrial bioenergetic function, intracellular ATP levels, intracellular and tissue concentrations of tested compounds, and in vivo tumor growth were performed. Both Mito-ChM and Mito-ChMAc selectively depleted intracellular ATP and caused prolonged inhibition of ATP-linked oxygen consumption rate in breast cancer cells, but not in non-cancerous cells. These effects were significantly augmented by inhibition of glycolysis. Mito-ChM and Mito-ChMAc exhibited anti-proliferative effects and cytotoxicity in several breast cancer cells with different genetic background. Furthermore, Mito-ChM selectively accumulated in tumor tissue and inhibited tumor growth in a xenograft model of human breast cancer. We conclude that mitochondria-targeted small molecular weight chromanols exhibit selective anti-proliferative effects and cytotoxicity in multiple breast cancer cells, and that esterification of the hydroxyl group in mito-chromanols is not a critical requirement for its anti-proliferative and cytotoxic effect.

  10. Mitochondria-targeted vitamin E analogs inhibit breast cancer cell energy metabolism and promote cell death

    PubMed Central

    2013-01-01

    Background Recent research has revealed that targeting mitochondrial bioenergetic metabolism is a promising chemotherapeutic strategy. Key to successful implementation of this chemotherapeutic strategy is the use of new and improved mitochondria-targeted cationic agents that selectively inhibit energy metabolism in breast cancer cells, while exerting little or no long-term cytotoxic effect in normal cells. Methods In this study, we investigated the cytotoxicity and alterations in bioenergetic metabolism induced by mitochondria-targeted vitamin E analog (Mito-chromanol, Mito-ChM) and its acetylated ester analog (Mito-ChMAc). Assays of cell death, colony formation, mitochondrial bioenergetic function, intracellular ATP levels, intracellular and tissue concentrations of tested compounds, and in vivo tumor growth were performed. Results Both Mito-ChM and Mito-ChMAc selectively depleted intracellular ATP and caused prolonged inhibition of ATP-linked oxygen consumption rate in breast cancer cells, but not in non-cancerous cells. These effects were significantly augmented by inhibition of glycolysis. Mito-ChM and Mito-ChMAc exhibited anti-proliferative effects and cytotoxicity in several breast cancer cells with different genetic background. Furthermore, Mito-ChM selectively accumulated in tumor tissue and inhibited tumor growth in a xenograft model of human breast cancer. Conclusions We conclude that mitochondria-targeted small molecular weight chromanols exhibit selective anti-proliferative effects and cytotoxicity in multiple breast cancer cells, and that esterification of the hydroxyl group in mito-chromanols is not a critical requirement for its anti-proliferative and cytotoxic effect. PMID:23764021

  11. A novel bifunctional mitochondria-targeted anticancer agent with high selectivity for cancer cells

    PubMed Central

    He, Huan; Li, Dong-Wei; Yang, Li-Yun; Fu, Li; Zhu, Xun-Jin; Wong, Wai-Kwok; Jiang, Feng-Lei; Liu, Yi

    2015-01-01

    Mitochondria have recently emerged as novel targets for cancer therapy due to its important roles in fundamental cellular function. Discovery of new chemotherapeutic agents that allow for simultaneous treatment and visualization of cancer is urgent. Herein, we demonstrate a novel bifunctional mitochondria-targeted anticancer agent (FPB), exhibiting both imaging capability and anticancer activity. It can selectively accumulate in mitochondria and induce cell apoptosis. Notably, it results in much higher toxicity toward cancer cells owing to much higher uptake by cancer cells. These features make it highly attractive in cancer imaging and treatment. PMID:26337336

  12. Mitochondria-Targeted Antioxidant SkQ1 Improves Dermal Wound Healing in Genetically Diabetic Mice.

    PubMed

    Demyanenko, Ilya A; Zakharova, Vlada V; Ilyinskaya, Olga P; Vasilieva, Tamara V; Fedorov, Artem V; Manskikh, Vasily N; Zinovkin, Roman A; Pletjushkina, Olga Yu; Chernyak, Boris V; Skulachev, Vladimir P; Popova, Ekaterina N

    2017-01-01

    Oxidative stress is widely recognized as an important factor in the delayed wound healing in diabetes. However, the role of mitochondrial reactive oxygen species in this process is unknown. It was assumed that mitochondrial reactive oxygen species are involved in many wound-healing processes in both diabetic humans and animals. We have applied the mitochondria-targeted antioxidant 10-(6'-plastoquinonyl)decyltriphenylphosphonium (SkQ1) to explore the role of mitochondrial reactive oxygen species in the wound healing of genetically diabetic mice. Healing of full-thickness excisional dermal wounds in diabetic C57BL/KsJ-db(-)/db(-) mice was significantly enhanced after long-term (12 weeks) administration of SkQ1. SkQ1 accelerated wound closure and stimulated epithelization, granulation tissue formation, and vascularization. On the 7th day after wounding, SkQ1 treatment increased the number of α-smooth muscle actin-positive cells (myofibroblasts), reduced the number of neutrophils, and increased macrophage infiltration. SkQ1 lowered lipid peroxidation level but did not change the level of the circulatory IL-6 and TNF. SkQ1 pretreatment also stimulated cell migration in a scratch-wound assay in vitro under hyperglycemic condition. Thus, a mitochondria-targeted antioxidant normalized both inflammatory and regenerative phases of wound healing in diabetic mice. Our results pointed to nearly all the major steps of wound healing as the target of excessive mitochondrial reactive oxygen species production in type II diabetes.

  13. Mitochondria-Targeted Analogues of Metformin Exhibit Enhanced Antiproliferative and Radiosensitizing Effects in Pancreatic Cancer Cells.

    PubMed

    Cheng, Gang; Zielonka, Jacek; Ouari, Olivier; Lopez, Marcos; McAllister, Donna; Boyle, Kathleen; Barrios, Christy S; Weber, James J; Johnson, Bryon D; Hardy, Micael; Dwinell, Michael B; Kalyanaraman, Balaraman

    2016-07-01

    Metformin (Met) is an approved antidiabetic drug currently being explored for repurposing in cancer treatment based on recent evidence of its apparent chemopreventive properties. Met is weakly cationic and targets the mitochondria to induce cytotoxic effects in tumor cells, albeit not very effectively. We hypothesized that increasing its mitochondria-targeting potential by attaching a positively charged lipophilic substituent would enhance the antitumor activity of Met. In pursuit of this question, we synthesized a set of mitochondria-targeted Met analogues (Mito-Mets) with varying alkyl chain lengths containing a triphenylphosphonium cation (TPP(+)). In particular, the analogue Mito-Met10, synthesized by attaching TPP(+) to Met via a 10-carbon aliphatic side chain, was nearly 1,000 times more efficacious than Met at inhibiting cell proliferation in pancreatic ductal adenocarcinoma (PDAC). Notably, in PDAC cells, Mito-Met10 potently inhibited mitochondrial complex I, stimulating superoxide and AMPK activation, but had no effect in nontransformed control cells. Moreover, Mito-Met10 potently triggered G1 cell-cycle phase arrest in PDAC cells, enhanced their radiosensitivity, and more potently abrogated PDAC growth in preclinical mouse models, compared with Met. Collectively, our findings show how improving the mitochondrial targeting of Met enhances its anticancer activities, including aggressive cancers like PDAC in great need of more effective therapeutic options. Cancer Res; 76(13); 3904-15. ©2016 AACR. ©2016 American Association for Cancer Research.

  14. Mitochondria-Targeted Peptide Reverses Mitochondrial Dysfunction and Cognitive Deficits in Sepsis-Associated Encephalopathy.

    PubMed

    Wu, Jing; Zhang, Mingqiang; Hao, Shuangying; Jia, Ming; Ji, Muhuo; Qiu, Lili; Sun, Xiaoyan; Yang, Jianjun; Li, Kuanyu

    2015-08-01

    Sepsis-associated encephalopathy (SAE) is associated with increased mortality, morbidity, and long-term cognitive impairments. Its pathophysiology remains to be determined and an effective pharmacologic treatment is lacking. The goal of this study was to investigate the effects of the mitochondria-targeted peptide SS-31 on mitochondrial function and cognitive deficits in SAE mice. C57BL/6 male mice were randomly divided into sham, sham + SS-31, cecal ligation and puncture (CLP), and CLP + SS-31 groups. Peptide SS-31 (5 mg/kg) was intraperitoneally administrated immediately after operation and afterwards once daily for six consecutive days. Surviving mice were subjected to behavioral tests and the hippocampus was collected for biochemical analysis 7 days after operation. The results showed that CLP resulted in high mortality rate and cognitive deficits, representative characteristics of SAE. A physiological mechanistic investigation revealed that mitochondrial function of hippocampus was severely impaired, coupled with reactive oxygen species (ROS) generation, triggering neuronal apoptosis and inflammation. Notably, administration of peptide SS-31 protected the integrity of mitochondria, reversed the mitochondrial dysfunction, inhibited the apoptosis resulting from the release of cytochrome c, diminished the response of inflammation, and ultimately reversed the behavior deficits in the SAE mice. In conclusion, our data demonstrate that daily treatment with mitochondria-targeted peptide SS-31 reduces mortality rate and ameliorates cognitive deficits, which is possibly through a mechanism of reversing mitochondrial dysfunction and partial inhibition of neuronal apoptosis and inflammation in the hippocampus of the SAE mice.

  15. Mitochondria-targeted agents: Future perspectives of mitochondrial pharmaceutics in cardiovascular diseases

    PubMed Central

    Ajith, Thekkuttuparambil Ananthanarayanan; Jayakumar, Thankamani Gopinathan

    2014-01-01

    Mitochondria are one of the major sites for the generation of reactive oxygen species (ROS) as an undesirable side product of oxidative energy metabolism. Damaged mitochondria can augment the generation of ROS. Dysfunction of mitochondria increase the risk for a large number of human diseases, including cardiovascular diseases (CVDs). Heart failure (HF) following ischemic heart disease, infantile cardiomyopathy and cardiac hypertrophy associated with left ventricular dilations are some of the CVDs in which the role of mitochondrial oxidative stress has been reported. Advances in mitochondrial research during the last decade focused on the preservation of its function in the myocardium, which is vital for the cellular energy production. Experimental and clinical trials have been conducted using mitochondria-targeted molecules like: MnSOD mimetics, such as EUK-8, EUK-134 and MitoSOD; choline esters of glutathione and N-acetyl-L-cysteine; triphenylphosphonium ligated vitamin E, lipoic acid, plastoquinone and mitoCoQ10; and Szeto-Schiller (SS)- peptides (SS-02 and SS-31). Although many results are inconclusive, some of the findings, especially on CoQ10, are worthwhile. This review summarizes the role of mitochondria-targeted delivery of agents and their consequences in the control of HF. PMID:25349653

  16. Mitochondria-Targeted Triphenylphosphonium-Based Compounds: Syntheses, Mechanisms of Action, and Therapeutic and Diagnostic Applications.

    PubMed

    Zielonka, Jacek; Joseph, Joy; Sikora, Adam; Hardy, Micael; Ouari, Olivier; Vasquez-Vivar, Jeannette; Cheng, Gang; Lopez, Marcos; Kalyanaraman, Balaraman

    2017-08-09

    Mitochondria are recognized as one of the most important targets for new drug design in cancer, cardiovascular, and neurological diseases. Currently, the most effective way to deliver drugs specifically to mitochondria is by covalent linking a lipophilic cation such as an alkyltriphenylphosphonium moiety to a pharmacophore of interest. Other delocalized lipophilic cations, such as rhodamine, natural and synthetic mitochondria-targeting peptides, and nanoparticle vehicles, have also been used for mitochondrial delivery of small molecules. Depending on the approach used, and the cell and mitochondrial membrane potentials, more than 1000-fold higher mitochondrial concentration can be achieved. Mitochondrial targeting has been developed to study mitochondrial physiology and dysfunction and the interaction between mitochondria and other subcellular organelles and for treatment of a variety of diseases such as neurodegeneration and cancer. In this Review, we discuss efforts to target small-molecule compounds to mitochondria for probing mitochondria function, as diagnostic tools and potential therapeutics. We describe the physicochemical basis for mitochondrial accumulation of lipophilic cations, synthetic chemistry strategies to target compounds to mitochondria, mitochondrial probes, and sensors, and examples of mitochondrial targeting of bioactive compounds. Finally, we review published attempts to apply mitochondria-targeted agents for the treatment of cancer and neurodegenerative diseases.

  17. A Powerful Mitochondria-Targeted Iron Chelator Affords High Photoprotection against Solar Ultraviolet A Radiation.

    PubMed

    Reelfs, Olivier; Abbate, Vincenzo; Hider, Robert C; Pourzand, Charareh

    2016-08-01

    Mitochondria are the principal destination for labile iron, making these organelles particularly susceptible to oxidative damage on exposure to ultraviolet A (UVA, 320-400 nm), the oxidizing component of sunlight. The labile iron-mediated oxidative damage caused by UVA to mitochondria leads to necrotic cell death via adenosine triphosphate depletion. Therefore, targeted removal of mitochondrial labile iron via highly specific tools from these organelles may be an effective approach to protect the skin cells against the harmful effects of UVA. In this work, we designed a mitochondria-targeted hexadentate (tricatechol-based) iron chelator linked to mitochondria-homing SS-like peptides. The photoprotective potential of this compound against UVA-induced oxidative damage and cell death was evaluated in cultured primary skin fibroblasts. Our results show that this compound provides unprecedented protection against UVA-induced mitochondrial damage, adenosine triphosphate depletion, and the ensuing necrotic cell death in skin fibroblasts, and this effect is fully related to its potent iron-chelating property in the organelle. This mitochondria-targeted iron chelator has therefore promising potential for skin photoprotection against the deleterious effects of the UVA component of sunlight. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Treatment Strategies that Enhance the Efficacy and Selectivity of Mitochondria-Targeted Anticancer Agents

    PubMed Central

    Modica-Napolitano, Josephine S.; Weissig, Volkmar

    2015-01-01

    Nearly a century has passed since Otto Warburg first observed high rates of aerobic glycolysis in a variety of tumor cell types and suggested that this phenomenon might be due to an impaired mitochondrial respiratory capacity in these cells. Subsequently, much has been written about the role of mitochondria in the initiation and/or progression of various forms of cancer, and the possibility of exploiting differences in mitochondrial structure and function between normal and malignant cells as targets for cancer chemotherapy. A number of mitochondria-targeted compounds have shown efficacy in selective cancer cell killing in pre-clinical and early clinical testing, including those that induce mitochondria permeability transition and apoptosis, metabolic inhibitors, and ROS regulators. To date, however, none has exhibited the standards for high selectivity and efficacy and low toxicity necessary to progress beyond phase III clinical trials and be used as a viable, single modality treatment option for human cancers. This review explores alternative treatment strategies that have been shown to enhance the efficacy and selectivity of mitochondria-targeted anticancer agents in vitro and in vivo, and may yet fulfill the clinical promise of exploiting the mitochondrion as a target for cancer chemotherapy. PMID:26230693

  19. Mitochondria-Targeted Antioxidant Prevents Cardiac Dysfunction Induced by Tafazzin Gene Knockdown in Cardiac Myocytes

    PubMed Central

    He, Quan; Harris, Nicole; Ren, Jun; Han, Xianlin

    2014-01-01

    Tafazzin, a mitochondrial acyltransferase, plays an important role in cardiolipin side chain remodeling. Previous studies have shown that dysfunction of tafazzin reduces cardiolipin content, impairs mitochondrial function, and causes dilated cardiomyopathy in Barth syndrome. Reactive oxygen species (ROS) have been implicated in the development of cardiomyopathy and are also the obligated byproducts of mitochondria. We hypothesized that tafazzin knockdown increases ROS production from mitochondria, and a mitochondria-targeted antioxidant prevents tafazzin knockdown induced mitochondrial and cardiac dysfunction. We employed cardiac myocytes transduced with an adenovirus containing tafazzin shRNA as a model to investigate the effects of the mitochondrial antioxidant, mito-Tempo. Knocking down tafazzin decreased steady state levels of cardiolipin and increased mitochondrial ROS. Treatment of cardiac myocytes with mito-Tempo normalized tafazzin knockdown enhanced mitochondrial ROS production and cellular ATP decline. Mito-Tempo also significantly abrogated tafazzin knockdown induced cardiac hypertrophy, contractile dysfunction, and cell death. We conclude that mitochondria-targeted antioxidant prevents cardiac dysfunction induced by tafazzin gene knockdown in cardiac myocytes and suggest mito-Tempo as a potential therapeutic for Barth syndrome and other dilated cardiomyopathies resulting from mitochondrial oxidative stress. PMID:25247053

  20. Mitochondria-targeted antioxidants as promising drugs for treatment of age-related brain diseases.

    PubMed

    Skulachev, Vladimir P

    2012-01-01

    Much experimental evidence suggests that age-related brain pathologies are most often mediated by reactive oxygen species primarily originating from mitochondria (mROS). Two papers with such evidence have been recently published in the Journal of Alzheimer's Disease (Stefanova et al., J Alzheimers Dis 21, 476-491, 2010; Lloret et al., J Alzheimers Dis, doi: 10.3233/JAD-2011-110890). In the first paper, it was shown that appearance of a typical behavioral trait of aging in rats (that old animals do not enter an open arm in a maze) was completely reversed by ten weeks treatment of the old rats with the mitochondria-targeted antioxidant SkQ1. In the second article, the authors identified molecular mechanisms by which amyloid-β-induced mROS can mediate hyperphosphorylation of the tau protein, a key event in Alzheimer's disease. Conventional antioxidants prevented such hyperphosphorylation. In this article, I will summarize the present state of the art in this field. I conclude that mitochondria-targeted rechargeable antioxidants are promising as tools to treat brain pathologies developing in elderly humans.

  1. In search of novel highly active mitochondria-targeted antioxidants: thymoquinone and its cationic derivatives.

    PubMed

    Severina, Inna I; Severin, Fedor F; Korshunova, Galina A; Sumbatyan, Natalya V; Ilyasova, Tatyana M; Simonyan, Ruben A; Rogov, Anton G; Trendeleva, Tatyana A; Zvyagilskaya, Renata A; Dugina, Vera B; Domnina, Lidia V; Fetisova, Elena K; Lyamzaev, Konstantin G; Vyssokikh, Mikhail Yu; Chernyak, Boris V; Skulachev, Maxim V; Skulachev, Vladimir P; Sadovnichii, Viktor A

    2013-06-27

    Since the times of the Bible, an extract of black cumin seeds was used as a medicine to treat many human pathologies. Thymoquinone (2-demethylplastoquinone derivative) was identified as an active antioxidant component of this extract. Recently, it was shown that conjugates of plastoquinone and penetrating cations are potent mitochondria-targeted antioxidants effective in treating a large number of age-related pathologies. This review summarizes new data on the antioxidant and some other properties of membrane-penetrating cationic compounds where 2-demethylplastoquinone substitutes for plastoquinone. It was found that such a substitution significantly increases a window between anti- and prooxidant concentrations of the conjugates. Like the original plastoquinone derivatives, the novel compounds are easily reduced by the respiratory chain, penetrate through model and natural membranes, specifically accumulate in mitochondria in an electrophoretic fashion, and strongly inhibit H2O2-induced apoptosis at pico- and nanomolar concentrations in cell cultures. At present, cationic demethylplastoquinone derivatives appear to be the most promising mitochondria-targeted drugs of the quinone series.

  2. Tumor mitochondria-targeted photodynamic therapy with a translocator protein (TSPO)-specific photosensitizer.

    PubMed

    Zhang, Shaojuan; Yang, Ling; Ling, Xiaoxi; Shao, Pin; Wang, Xiaolei; Edwards, W Barry; Bai, Mingfeng

    2015-12-01

    Photodynamic therapy (PDT) has been proven to be a minimally invasive and effective therapeutic strategy for cancer treatment. It can be used alone or as a complement to conventional cancer treatments, such as surgical debulking and chemotherapy. The mitochondrion is an attractive target for developing novel PDT agents, as it produces energy for cells and regulates apoptosis. Current strategy of mitochondria targeting is mainly focused on utilizing cationic photosensitizers that bind to the negatively charged mitochondria membrane. However, such an approach is lack of selectivity of tumor cells. To minimize the damage on healthy tissues and improve therapeutic efficacy, an alternative targeting strategy with high tumor specificity is in critical need. Herein, we report a tumor mitochondria-specific PDT agent, IR700DX-6T, which targets the 18kDa mitochondrial translocator protein (TSPO). IR700DX-6T induced apoptotic cell death in TSPO-positive breast cancer cells (MDA-MB-231) but not TSPO-negative breast cancer cells (MCF-7). In vivo PDT study suggested that IR700DX-6T-mediated PDT significantly inhibited the growth of MDA-MB-231 tumors in a target-specific manner. These combined data suggest that this new TSPO-targeted photosensitizer has great potential in cancer treatment. Photodynamic therapy (PDT) is an effective and minimally invasive therapeutic technique for treating cancers. Mitochondrion is an attractive target for developing novel PDT agents, as it produces energy to cells and regulates apoptosis. Current mitochondria targeted photosensitizers (PSs) are based on cationic molecules, which interact with the negatively charged mitochondria membrane. However, such PSs are not specific for cancerous cells, which may result in unwanted side effects. In this study, we developed a tumor mitochondria-targeted PS, IR700DX-6T, which binds to translocator protein (TSPO). This agent effectively induced apoptosis in TSPO-positive cancer cells and significantly

  3. Mitochondria-targeted liposomes improve the apoptotic and cytotoxic action of sclareol.

    PubMed

    Patel, Niravkumar R; Hatziantoniou, Sophia; Georgopoulos, Aristidis; Demetzos, Costas; Torchilin, Vladimir P; Weissig, Volkmar; D'Souza, Gerard G M

    2010-09-01

    Current efforts toward improving the effectiveness of drug therapy are increasingly relying on drug-targeting strategies to effectively deliver bioactive molecules to their molecular targets. Pharmaceutical nanocarriers represent a major tool toward this aim, and our efforts have been directed toward achieving nanocarrier-mediated subcellular delivery of drug molecules with mitochondria as the primary subcellular target. Meeting the need for specific subcellular delivery is essential to realizing the full potential of many poorly soluble anticancer drugs. In this article, we report that mitochondria-targeted liposomes significantly improve the apoptotic and cytotoxic action of sclareol, a poorly soluble potential anticancer drug. The results support the broad applicability of our nanocarrier-mediated subcellular targeting approach as a means to improve the effectiveness of certain anticancer therapeutics.

  4. A mitochondria-targeted ratiometric two-photon fluorescent probe for biological zinc ions detection

    PubMed Central

    Ning, Peng; Jiang, Jiacheng; Li, Longchun; Wang, Shuxin; Yu, Haizhu; Feng, Yan; Zhu, Manzhou; Zhang, Buchang; Yin, Hang; Guo, Qingxiang; Meng, Xiangming

    2015-01-01

    A mitochondria-targeted ratiometric two-photon fluorescent probe (Mito-MPVQ) for biological zinc ions detection was developed based on quinolone platform. Mito-MPVQ showed large red shifts (68nm) and selective ratiometric signal upon Zn2+ binding. The ratio of emission intensity (I488 nm/I420 nm) increases dramatically from 0.45 to 3.79 (ca. 8-fold). NMR titration and theoretical calculation confirmed the binding of Mito-MPVQ and Zn2+. Mito-MPVQ also exhibited large two-photon absorption cross sections (150GM) at nearly 720 nm and insensitivity to pH within the biologically relevant pH range. Cell imaging indicated that Mito-MPVQ could efficiently located in mitochondria and monitor mitochondrial Zn2+ under two-photon excitation with low cytotoxicity. PMID:26528806

  5. The Use of Mitochondria-Targeted Endonucleases to Manipulate mtDNA

    PubMed Central

    Bacman, Sandra R.; Williams, Sion L.; Pinto, Milena; Moraes, Carlos T.

    2014-01-01

    For more than a decade, mitochondria-targeted nucleases have been used to promote double-strand breaks in the mitochondrial genome. This was done in mitochondrial DNA (mtDNA) homoplasmic systems, where all mtDNA molecules can be affected, to create models of mitochondrial deficiencies. Alternatively, they were also used in a heteroplasmic model, where only a subset of the mtDNA molecules were substrates for cleavage. The latter approach showed that mitochondrial-targeted nucleases can reduce mtDNA haplotype loads in affected tissues, with clear implications for the treatment of patients with mitochondrial diseases. In the last few years, designer nucleases, such as ZFN and TALEN, have been adapted to cleave mtDNA, greatly expanding the potential therapeutic use. This chapter describes the techniques and approaches used to test these designer enzymes. PMID:25416366

  6. Mitochondria-targeted antioxidant Mito-Tempo protects against acetaminophen hepatotoxicity.

    PubMed

    Du, Kuo; Farhood, Anwar; Jaeschke, Hartmut

    2017-02-01

    Acetaminophen (APAP) hepatotoxicity is characterized by an extensive mitochondrial oxidant stress. However, its importance as a drug target has not been clarified. To investigate this, fasted C57BL/6J mice were treated with 300 mg/kg APAP and the mitochondria-targeted antioxidant Mito-Tempo (MT) was given 1.5 h later. APAP caused severe liver injury in mice, as indicated by the increase in plasma ALT activities and centrilobular necrosis. MT dose-dependently reduced the injury. Importantly, MT did not affect APAP-protein adducts formation, glutathione depletion or c-jun N-terminal kinase activation and its mitochondrial translocation. In contrast, hepatic glutathione disulfide and peroxynitrite formation were dose-dependently reduced by MT, indicating its effective mitochondrial oxidant stress scavenging capacity. Consequently, mitochondrial translocation of Bax and release of mitochondrial intermembrane proteins such as apoptosis-inducing factor were prevented, and nuclear DNA fragmentation was eliminated. To demonstrate the importance of mitochondria-specific antioxidant property of MT, we compared its efficacy with Tempo, which has the same pharmacological mode of action as MT but lacks the mitochondria targeting moiety. In contrast to the dramatic protection by MT, the same molar dose of Tempo did not significantly reduce APAP hepatotoxicity. In contrast, even a 3 h post-treatment with MT reduced 70 % of the injury, and the combination of MT with N-acetylcysteine (NAC) provided superior protection than NAC alone. We conclude that MT protects against APAP overdose in mice by attenuating the mitochondrial oxidant stress and preventing peroxynitrite formation and the subsequent mitochondrial dysfunction. MT is a promising therapeutic agent for APAP overdose patients.

  7. Mitochondria-targeted Antioxidants as a Prospective Therapeutic Strategy for Multiple Sclerosis.

    PubMed

    Fetisova, Elena; Chernyak, Boris; Korshunova, Galina; Muntyan, Maria; Skulachev, Vladimir

    2017-01-01

    Multiple sclerosis (MS) is one of the most widespread chronic neurological diseases that manifests itself by progressive demyelination in the central nervous system. The study of MS pathogenesis begins with the onset of the relapsing-remitting phase of the disease, which becomes apparent due to microglia activation, neuroinflammation and demyelination/ remyelination in the white matter. The following progressive phase is accompanied by severe neurological symptoms when demyelination and neurodegeneration are spread to both gray and white matter. In this review, we discuss a possible role of mitochondrial reactive oxygen species (mtROS) in MS pathogenesis, mechanisms of mtROS generation and effects of some mitochondria-targeted antioxidants as potential components of MS therapy. In the early phase of MS, mtROS stimulate NLRP3 inflammasomes, which is critical for the formation of local inflammatory lesions. Later, mtROS contribute to blood-brain barrier disruption induced by mediators of inflammation, followed by infiltration of leukocytes. ROS generated by leukocytes and activated microglia promote mitochondrial dysfunction and oligodendrocyte cell death. In the progressive phase, neurodegeneration also depends on excessive mtROS generation. Currently, only a few immunomodulatory drugs are approved for treatment of MS. These drugs mainly reduce the number of relapses but do not stop MS progression. Certain dietary and synthetic antioxidants have demonstrated encouraging results in animal models of MS but were ineffective in the completed clinical trials. Novel mitochondria-targeted antioxidants could be promising components of combined programs for MS therapy considering that they can be applied at extremely low doses and concurrently demonstrate anti-inflammatory and neuroprotective activities. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  8. Pathophysiological and pharmacological implications of mitochondria-targeted reactive oxygen species generation in astrocytes.

    PubMed

    Jou, Mei-Jie

    2008-01-01

    Astrocytes, in addition to passively supporting neurons, have recently been shown to be actively involved in synaptic transmission and neurovascular coupling in the central nervous system (CNS). This review summarizes briefly our previous observations using fluorescent probes coupled with laser scanning digital imaging microscopy to visualize spatio-temporal alteration of mitochondrial reactive oxygen species (mROS) generation in intact astrocytes. mROS formation is enhanced by exogenous oxidants exposure, Ca2+ stress and endogenous pathological defect of mitochondrial respiratory complexes. In addition, mROS formation can be specifically stimulated by visible light or visible laser irradiation and can be augmented further by photodynamic coupling with photosensitizers, particularly with mitochondria-targeted photosensitizers. "Severe" oxidative insult often results in massive and homogeneous augmentation of mROS formation which causes cessation of mitochondrial movement, pathological fission and irreversible swelling of mitochondria and eventually apoptosis or necrosis of cells. Mitochondria-targeted antioxidants and protectors such as MitoQ, melatonin and nanoparticle C(60) effectively prevent "severe" mROS generation. Intriguingly, "minor" oxidative insults enhance heterogeneity of mROS and mitochondrial dynamics. "Minor" mROS formation-induced fission and fusion of mitochondria relocates mitochondrial network to form a mitochondria free gap, i.e., "firewall", which may play a crucial role in mROS-mediated protective "preconditioning" by preventing propagation of mROS during oxidative insults. These mROS-targeted strategies for either enhancement or prevention of mitochondrial oxidative stress in astrocytes may provide new insights for future development of therapeutic interventions in the treatment of cancer such as astrocytomas and gliomas and astrocyte-associated neurodegeneration, mitochondrial diseases and aging.

  9. Mitochondria-targeted antioxidant SkQ1 improves impaired dermal wound healing in old mice.

    PubMed

    Demyanenko, Ilya A; Popova, Ekaterina N; Zakharova, Vlada V; Ilyinskaya, Olga P; Vasilieva, Tamara V; Romashchenko, Valeria P; Fedorov, Artem V; Manskikh, Vasily N; Skulachev, Maxim V; Zinovkin, Roman A; Pletjushkina, Olga Yu; Skulachev, Vladimir P; Chernyak, Boris V

    2015-07-01

    The process of skin wound healing is delayed or impaired in aging animals. To investigate the possible role of mitochondrial reactive oxygen species (mtROS) in cutaneous wound healing of aged mice, we have applied the mitochondria-targeted antioxidant SkQ1. The SkQ1 treatment resulted in accelerated resolution of the inflammatory phase, formation of granulation tissue, vascularization and epithelization of the wounds. The wounds of SkQ1-treated mice contained increased amount of myofibroblasts which produce extracellular matrix proteins and growth factors mediating granulation tissue formation. This effect resembled SkQ1-induced differentiation of fibroblasts to myofibroblast, observed earlierin vitro. The Transforming Growth Factor beta (TGFb) produced by SkQ1-treated fibroblasts was found to stimulated motility of endothelial cells in vitro, an effect which may underlie pro-angiogenic action of SkQ1 in the wounds. In vitro experiments showed that SkQ1 prevented decomposition of VE-cadherin containing contacts and following increase in permeability of endothelial cells monolayer, induced by pro-inflammatory cytokine TNF. Prevention of excessive reaction of endothelium to the pro-inflammatory cytokine(s) might account for anti-inflammatory effect of SkQ1. Our findings point to an important role of mtROS in pathogenesis of age-related chronic wounds.

  10. Mitochondria targeted peptides protect against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity.

    PubMed

    Yang, Lichuan; Zhao, Kesheng; Calingasan, Noel Y; Luo, Guoxiong; Szeto, Hazel H; Beal, M Flint

    2009-09-01

    A large body of evidence suggests that mitochondrial dysfunction and oxidative damage play a role in the pathogenesis of Parkinson's disease (PD). A number of antioxidants have been effective in animal models of PD. We have developed a family of mitochondria-targeted peptides that can protect against mitochondrial swelling and apoptosis (SS peptides). In this study, we examined the ability of two peptides, SS-31 and SS-20, to protect against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxicity in mice. SS-31 produced dose-dependent complete protection against loss of dopamine and its metabolites in striatum, as well as loss of tyrosine hydroxylase immunoreactive neurons in substantia nigra pars compacta. SS-20, which does not possess intrinsic ability in scavenging reactive oxygen species, also demonstrated significant neuroprotective effects on dopaminergic neurons of MPTP-treated mice. Both SS-31 and SS-20 were very potent (nM) in preventing MPP+ (1-methyl-4-phenylpyridinium)-induced cell death in cultured dopamine cells (SN4741). Studies with isolated mitochondria showed that both SS-31 and SS-20 prevented MPP+-induced inhibition of oxygen consumption and ATP production, and mitochondrial swelling. These findings provide strong evidence that these neuroprotective peptides, which target both mitochondrial dysfunction and oxidative damage, are a promising approach for the treatment of PD.

  11. Mitochondria Targeted Peptides Protect Against 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Neurotoxicity

    PubMed Central

    Yang, Lichuan; Zhao, Kesheng; Calingasan, Noel Y.; Luo, Guoxiong; Szeto, Hazel H.

    2009-01-01

    Abstract A large body of evidence suggests that mitochondrial dysfunction and oxidative damage play a role in the pathogenesis of Parkinson's disease (PD). A number of antioxidants have been effective in animal models of PD. We have developed a family of mitochondria-targeted peptides that can protect against mitochondrial swelling and apoptosis (SS peptides). In this study, we examined the ability of two peptides, SS-31 and SS-20, to protect against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxicity in mice. SS-31 produced dose-dependent complete protection against loss of dopamine and its metabolites in striatum, as well as loss of tyrosine hydroxylase immunoreactive neurons in substantia nigra pars compacta. SS-20, which does not possess intrinsic ability in scavenging reactive oxygen species, also demonstrated significant neuroprotective effects on dopaminergic neurons of MPTP-treated mice. Both SS-31 and SS-20 were very potent (nM) in preventing MPP+ (1-methyl-4-phenylpyridinium)-induced cell death in cultured dopamine cells (SN4741). Studies with isolated mitochondria showed that both SS-31 and SS-20 prevented MPP+-induced inhibition of oxygen consumption and ATP production, and mitochondrial swelling. These findings provide strong evidence that these neuroprotective peptides, which target both mitochondrial dysfunction and oxidative damage, are a promising approach for the treatment of PD. Antioxid. Redox Signal. 11, 2095–2104. PMID:19203217

  12. A mitochondria-targeted mass spectrometry probe to detect glyoxals: implications for diabetes☆

    PubMed Central

    Pun, Pamela Boon Li; Logan, Angela; Darley-Usmar, Victor; Chacko, Balu; Johnson, Michelle S.; Huang, Guang W.; Rogatti, Sebastian; Prime, Tracy A.; Methner, Carmen; Krieg, Thomas; Fearnley, Ian M.; Larsen, Lesley; Larsen, David S.; Menger, Katja E.; Collins, Yvonne; James, Andrew M.; Kumar, G.D. Kishore; Hartley, Richard C.; Smith, Robin A.J.; Murphy, Michael P.

    2014-01-01

    The glycation of protein and nucleic acids that occurs as a consequence of hyperglycemia disrupts cell function and contributes to many pathologies, including those associated with diabetes and aging. Intracellular glycation occurs after the generation of the reactive 1,2-dicarbonyls methylglyoxal and glyoxal, and disruption of mitochondrial function is associated with hyperglycemia. However, the contribution of these reactive dicarbonyls to mitochondrial damage in pathology is unclear owing to uncertainties about their levels within mitochondria in cells and in vivo. To address this we have developed a mitochondria-targeted reagent (MitoG) designed to assess the levels of mitochondrial dicarbonyls within cells. MitoG comprises a lipophilic triphenylphosphonium cationic function, which directs the molecules to mitochondria within cells, and an o-phenylenediamine moiety that reacts with dicarbonyls to give distinctive and stable products. The extent of accumulation of these diagnostic heterocyclic products can be readily and sensitively quantified by liquid chromatography–tandem mass spectrometry, enabling changes to be determined. Using the MitoG-based analysis we assessed the formation of methylglyoxal and glyoxal in response to hyperglycemia in cells in culture and in the Akita mouse model of diabetes in vivo. These findings indicated that the levels of methylglyoxal and glyoxal within mitochondria increase during hyperglycemia both in cells and in vivo, suggesting that they can contribute to the pathological mitochondrial dysfunction that occurs in diabetes and aging. PMID:24316194

  13. Effects of the Mitochondria-Targeted Antioxidant Mitoquinone in Murine Acute Pancreatitis

    PubMed Central

    Wen, Li; Szatmary, Peter; Mukherjee, Rajarshi; Armstrong, Jane; Chvanov, Michael; Tepikin, Alexei V.; Murphy, Michael P.; Sutton, Robert; Criddle, David N.

    2015-01-01

    Although oxidative stress has been strongly implicated in the development of acute pancreatitis (AP), antioxidant therapy in patients has so far been discouraging. The aim of this study was to assess potential protective effects of a mitochondria-targeted antioxidant, MitoQ, in experimental AP using in vitro and in vivo approaches. MitoQ blocked H2O2-induced intracellular ROS responses in murine pancreatic acinar cells, an action not shared by the control analogue dTPP. MitoQ did not reduce mitochondrial depolarisation induced by either cholecystokinin (CCK) or bile acid TLCS, and at 10 µM caused depolarisation per se. Both MitoQ and dTPP increased basal and CCK-induced cell death in a plate-reader assay. In a TLCS-induced AP model MitoQ treatment was not protective. In AP induced by caerulein hyperstimulation (CER-AP), MitoQ exerted mixed effects. Thus, partial amelioration of histopathology scores was observed, actions shared by dTPP, but without reduction of the biochemical markers pancreatic trypsin or serum amylase. Interestingly, lung myeloperoxidase and interleukin-6 were concurrently increased by MitoQ in CER-AP. MitoQ caused biphasic effects on ROS production in isolated polymorphonuclear leukocytes, inhibiting an acute increase but elevating later levels. Our results suggest that MitoQ would be inappropriate for AP therapy, consistent with prior antioxidant evaluations in this disease. PMID:25878403

  14. Perspectives and potential applications of mitochondria-targeted antioxidants in cardiometabolic diseases and type 2 diabetes.

    PubMed

    Rocha, Milagros; Apostolova, Nadezda; Herance, Jose Raul; Rovira-Llopis, Susana; Hernandez-Mijares, Antonio; Victor, Victor M

    2014-01-01

    There is abundant evidence to suggest that mitochondrial dysfunction is a main cause of insulin resistance and related cardiometabolic comorbidities. On the other hand, insulin resistance is one of the main characteristics of type 2 diabetes, obesity, and metabolic syndrome. Lipid and glucose metabolism require mitochondria to generate energy, and when O2 consumption is low due to inefficient nutrient oxidation, there is an increase in reactive oxygen species, which can impair different types of molecules, including DNA, lipids, proteins, and carbohydrates, thereby inducing proinflammatory processes. Factors which contribute to mitochondrial dysfunction, such as mitochondrial biogenesis and genetics, can also lead to insulin resistance in different insulin-target tissues, and its association with mitochondrial dysfunction can culminate in the development of cardiovascular diseases. In this context, therapies that improve mitochondrial function may also improve insulin resistance. This review explains mechanisms of mitochondrial function related to the pathological effects of insulin resistance in different tissues. The pathogenesis of cardiometabolic diseases will be explained from a mitochondrial perspective and the potential beneficial effects of mitochondria-targeted antioxidants as a therapy for modulating mitochondrial function in cardiometabolic diseases, especially diabetes, will also be considered.

  15. Vandetanib and cabozantinib potentiate mitochondria-targeted agents to suppress medullary thyroid carcinoma cells.

    PubMed

    Starenki, Dmytro; Hong, Seung-Keun; Wu, Pui-Kei; Park, Jong-In

    2017-07-03

    Although the FDA-approved receptor tyrosine kinases inhibitors, vandetanib and cabozantinib, are used to treat surgically inoperable progressive medullary thyroid carcinoma (MTC), not all patients are responsive while the disease sometimes progresses after an initial response. To better understand MTC drug resistance at molecular and biochemical levels, we have generated drug-resistant subpopulations of the human MTC cell lines, TT and MZ-CRC-1, via prolonged exposure to vandetanib and cabozantinib. These drug-resistant progenies exhibited substantial cross-resistance to vandetanib and cabozantinib, suggesting that these inhibitors may invoke an overlapping resistance mechanism(s) in MTC cells. Of note, vandetanib and cabozantinib increased mitochondrial membrane potential (Δψm) in drug-naïve as well as drug-resistant cells but only drug-naïve cells exhibited substantially altered oxygen consumption and extracellular acidification rates. Therefore, these inhibitors appear to cause a bioenergetics stress to which drug-resistant MTC cells are more tolerant. Given the ability of vandetanib and cabozantinib to increase Δψm, we hypothesized that these inhibitors can augment growth inhibitory effects of mitochondria-targeted carboxy-proxyl and ubiquinone by increasing their Δψm-dependent uptake/retention in MTC cells. Indeed, our in vitro and mouse xenograft data strongly support this possibility.

  16. Neuroprotection by a mitochondria-targeted drug in a Parkinson’s disease model

    PubMed Central

    Ghosh, Anamitra; Chandran, Karunakaran; Kalivendi, Shasi V.; Joseph, Joy; Antholine, William E.; Hillard, Cecilia J.; Kanthasamy, Arthi; Kanthasamy, Anumantha; Kalyanaraman, Balaraman

    2014-01-01

    The objective of this study was to assess the neuroprotective effects of a mitochondria-targeted antioxidant, Mito-Q10, the coenzyme-Q analog attached to a triphenylphosphonium cation that targets the antioxidant to mitochondria, in experimental models of Parkinson’s disease (PD). Primary mesencephalic neuronal cells and cultured dopaminergic cells were treated with 1-methyl-4-phenylpyridinium (MPP+), an active metabolite of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and mice were used for testing the efficacy of Mito-Q10. MPP+ treatment caused a dose-dependent loss of tyrosine hydroxylase and membrane potential and an increase in caspase-3 activation in dopaminergic cells, which were reversed by Mito-Q10. MPTP treatment induced a loss of striatal dopamine and its metabolites, inactivation of mitochondrial aconitase in the substantia nigra, and a loss of locomotor activity in mice. Treatment with Mito-Q10 significantly inhibited both MPP+- and MPTP-induced neurotoxicity in cell culture and mouse models. Collectively, these results indicate that mitochondrial targeting of antioxidants is a promising neuroprotective strategy in this preclinical mouse model of PD. PMID:20828611

  17. Mitochondria-targeted antioxidants and metabolic modulators as pharmacological interventions to slow ageing.

    PubMed

    Gruber, Jan; Fong, Sheng; Chen, Ce-Belle; Yoong, Sialee; Pastorin, Giorgia; Schaffer, Sebastian; Cheah, Irwin; Halliwell, Barry

    2013-01-01

    Populations in many nations today are rapidly ageing. This unprecedented demographic change represents one of the main challenges of our time. A defining property of the ageing process is a marked increase in the risk of mortality and morbidity with age. The incidence of cancer, cardiovascular and neurodegenerative diseases increases non-linearly, sometimes exponentially with age. One of the most important tasks in biogerontology is to develop interventions leading to an increase in healthy lifespan (health span), and a better understanding of basic mechanisms underlying the ageing process itself may lead to interventions able to delay or prevent many or even all age-dependent conditions. One of the putative basic mechanisms of ageing is age-dependent mitochondrial deterioration, closely associated with damage mediated by reactive oxygen species (ROS). Given the central role that mitochondria and mitochondrial dysfunction play not only in ageing but also in apoptosis, cancer, neurodegeneration and other age-related diseases there is great interest in approaches to protect mitochondria from ROS-mediated damage. In this review, we explore strategies of targeting mitochondria to reduce mitochondrial oxidative damage with the aim of preventing or delaying age-dependent decline in mitochondrial function and some of the resulting pathologies. We discuss mitochondria-targeted and -localized antioxidants (e.g.: MitoQ, SkQ, ergothioneine), mitochondrial metabolic modulators (e.g. dichloroacetic acid), and uncouplers (e.g.: uncoupling proteins, dinitrophenol) as well as some alternative future approaches for targeting compounds to the mitochondria, including advances from nanotechnology.

  18. Structural modifications of mitochondria-targeted chlorambucil alter cell death mechanism but preserve MDR evasion.

    PubMed

    Jean, Sae Rin; Pereira, Mark P; Kelley, Shana O

    2014-08-04

    Multidrug resistance (MDR) remains one of the major obstacles in chemotherapy, potentially rendering a multitude of drugs ineffective. Previously, we have demonstrated that mitochondrial targeting of DNA damaging agents is a promising tool for evading a number of common resistance factors that are present in the nucleus or cytosol. In particular, mitochondria-targeted chlorambucil (mt-Cbl) has increased potency and activity against resistant cancer cells compared to the parent compound chlorambucil (Cbl). However, it was found that, due to its high reactivity, mt-Cbl induces a necrotic type of cell death via rapid nonspecific alkylation of mitochondrial proteins. Here, we demonstrate that by tuning the alkylating activity of mt-Cbl via chemical modification, the rate of generation of protein adducts can be reduced, resulting in a shift of the cell death mechanism from necrosis to a more controlled apoptotic pathway. Moreover, we demonstrate that all of the modified mt-Cbl compounds effectively evade MDR resulting from cytosolic GST-μ upregulation by rapidly accumulating in mitochondria, inducing cell death directly from within. In this study, we systematically elucidated the advantages and limitations of targeting alkylating agents with varying reactivity to mitochondria.

  19. Measurement of Mitochondrial Cholesterol Import Using a Mitochondria-Targeted CYP11A1 Fusion Construct.

    PubMed

    Kennedy, Barry E; Charman, Mark; Karten, Barbara

    2017-01-01

    All animal membranes require cholesterol as an essential regulator of biophysical properties and function, but the levels of cholesterol vary widely among different subcellular compartments. Mitochondria, and in particular the inner mitochondrial membrane, have the lowest levels of cholesterol in the cell. Nevertheless, mitochondria need cholesterol for membrane maintenance and biogenesis, as well as oxysterol, steroid, and hepatic bile acid production. Alterations in mitochondrial cholesterol have been associated with a range of pathological conditions, including cancer, hepatosteatosis, cardiac ischemia, Alzheimer's, and Niemann-Pick Type C Disease. The mechanisms of mitochondrial cholesterol import are not fully elucidated yet, and may vary in different cell types and environmental conditions. Measuring cholesterol trafficking to the mitochondrial membranes is technically challenging because of its low abundance; for example, traditional pulse-chase experiments with isotope-labeled cholesterol are not feasible. Here, we describe improvements to a method first developed by the Miller group at the University of California to measure cholesterol trafficking to the inner mitochondrial membrane (IMM) through the conversion of cholesterol to pregnenolone. This method uses a mitochondria-targeted, ectopically expressed fusion construct of CYP11A1, ferredoxin reductase and ferredoxin. Pregnenolone is formed exclusively from cholesterol at the IMM, and can be analyzed with high sensitivity and specificity through ELISA or radioimmunoassay of the medium/buffer to reflect mitochondrial cholesterol import. This assay can be used to investigate the effects of genetic or pharmacological interventions on mitochondrial cholesterol import in cultured cells or isolated mitochondria.

  20. Mitochondria-targeted antioxidant preserves contractile properties and mitochondrial function of skeletal muscle in aged rats.

    PubMed

    Javadov, Sabzali; Jang, Sehwan; Rodriguez-Reyes, Natividad; Rodriguez-Zayas, Ana E; Soto Hernandez, Jessica; Krainz, Tanja; Wipf, Peter; Frontera, Walter

    2015-11-24

    Mitochondrial dysfunction plays a central role in the pathogenesis of sarcopenia associated with a loss of mass and activity of skeletal muscle. In addition to energy deprivation, increased mitochondrial ROS damage proteins and lipids in aged skeletal muscle. Therefore, prevention of mitochondrial ROS is important for potential therapeutic strategies to delay sarcopenia. This study elucidates the pharmacological efficiency of the new developed mitochondria-targeted ROS and electron scavenger, XJB-5-131 (XJB) to restore muscle contractility and mitochondrial function in aged skeletal muscle. Male adult (5-month old) and aged (29-month old) Fischer Brown Norway (F344/BN) rats were treated with XJB for four weeks and contractile properties of single skeletal muscle fibres and activity of mitochondrial ETC complexes were determined at the end of the treatment period. XJB-treated old rats showed higher muscle contractility associated with prevention of protein oxidation in both muscle homogenate and mitochondria compared with untreated counterparts. XJB-treated animals demonstrated a high activity of the respiratory complexes I, III, and IV with no changes in citrate synthase activity. These data demonstrate that mitochondrial ROS play a causal role in muscle weakness, and that a ROS scavenger specifically targeted to mitochondria can reverse age-related alterations of mitochondrial function and improve contractile properties in skeletal muscle.

  1. A mitochondria-targeted mass spectrometry probe to detect glyoxals: implications for diabetes.

    PubMed

    Pun, Pamela Boon Li; Logan, Angela; Darley-Usmar, Victor; Chacko, Balu; Johnson, Michelle S; Huang, Guang W; Rogatti, Sebastian; Prime, Tracy A; Methner, Carmen; Krieg, Thomas; Fearnley, Ian M; Larsen, Lesley; Larsen, David S; Menger, Katja E; Collins, Yvonne; James, Andrew M; Kumar, G D Kishore; Hartley, Richard C; Smith, Robin A J; Murphy, Michael P

    2014-02-01

    The glycation of protein and nucleic acids that occurs as a consequence of hyperglycemia disrupts cell function and contributes to many pathologies, including those associated with diabetes and aging. Intracellular glycation occurs after the generation of the reactive 1,2-dicarbonyls methylglyoxal and glyoxal, and disruption of mitochondrial function is associated with hyperglycemia. However, the contribution of these reactive dicarbonyls to mitochondrial damage in pathology is unclear owing to uncertainties about their levels within mitochondria in cells and in vivo. To address this we have developed a mitochondria-targeted reagent (MitoG) designed to assess the levels of mitochondrial dicarbonyls within cells. MitoG comprises a lipophilic triphenylphosphonium cationic function, which directs the molecules to mitochondria within cells, and an o-phenylenediamine moiety that reacts with dicarbonyls to give distinctive and stable products. The extent of accumulation of these diagnostic heterocyclic products can be readily and sensitively quantified by liquid chromatography-tandem mass spectrometry, enabling changes to be determined. Using the MitoG-based analysis we assessed the formation of methylglyoxal and glyoxal in response to hyperglycemia in cells in culture and in the Akita mouse model of diabetes in vivo. These findings indicated that the levels of methylglyoxal and glyoxal within mitochondria increase during hyperglycemia both in cells and in vivo, suggesting that they can contribute to the pathological mitochondrial dysfunction that occurs in diabetes and aging.

  2. Mitochondria Targeted and Intracellular Biothiol Triggered Hyperpolarized (129)Xe Magnetofluorescent Biosensor.

    PubMed

    Zeng, Qingbin; Guo, Qianni; Yuan, Yaping; Yang, Yuqi; Zhang, Bin; Ren, Lili; Zhang, Xiaoxiao; Luo, Qing; Liu, Maili; Bouchard, Louis-S; Zhou, Xin

    2017-02-21

    Biothiols such as gluthathione (GSH), cysteine (Cys), homocysteine (Hcy), and thioredoxin (Trx) play vital roles in cellular metabolism. Various diseases are associated with abnormal cellular biothiol levels. Thus, the intracellular detection of biothiol levels could be a useful diagnostic tool. A number of methods have been developed to detect intracellular thiols, but sensitivity and specificity problems have limited their applications. To address these limitations, we have designed a new biosensor based on hyperpolarized xenon magnetic resonance detection, which can be used to detect biothiol levels noninvasively. The biosensor is a multimodal probe that incorporates a cryptophane-A cage as (129)Xe NMR reporter, a naphthalimide moiety as fluorescence reporter, a disulfide bond as thiol-specific cleavable group, and a triphenylphosphonium moiety as mitochondria targeting unit. When the biosensor interacts with biothiols, disulfide bond cleavage leads to enhancements in the fluorescence intensity and changes in the (129)Xe chemical shift. Using Hyper-CEST (chemical exchange saturation transfer) NMR, our biosensor shows a low detection limit at picomolar (10(-10) M) concentration, which makes a promise to detect thiols in cells. The biosensor can detect biothiol effectively in live cells and shows good targeting ability to the mitochondria. This new approach not only offers a practical technique to detect thiols in live cells, but may also present an excellent in vivo test platform for xenon biosensors.

  3. Mito-DCA: a mitochondria targeted molecular scaffold for efficacious delivery of metabolic modulator dichloroacetate.

    PubMed

    Pathak, Rakesh K; Marrache, Sean; Harn, Donald A; Dhar, Shanta

    2014-05-16

    Tumor growth is fueled by the use of glycolysis, which normal cells use only in the scarcity of oxygen. Glycolysis makes tumor cells resistant to normal death processes. Targeting this unique tumor metabolism can provide an alternative strategy to selectively destroy the tumor, leaving normal tissue unharmed. The orphan drug dichloroacetate (DCA) is a mitochondrial kinase inhibitor that has the ability to show such characteristics. However, its molecular form shows poor uptake and bioavailability and limited ability to reach its target mitochondria. Here, we describe a targeted molecular scaffold for construction of a multiple DCA loaded compound, Mito-DCA, with three orders of magnitude enhanced potency and cancer cell specificity compared to DCA. Incorporation of a lipophilic triphenylphosphonium cation through a biodegradable linker in Mito-DCA allowed for mitochondria targeting. Mito-DCA did not show any significant metabolic effects toward normal cells but tumor cells with dysfunctional mitochondria were affected by Mito-DCA, which caused a switch from glycolysis to glucose oxidation and subsequent cell death via apoptosis. Effective delivery of DCA to the mitochondria resulted in significant reduction in lactate levels and played important roles in modulating dendritic cell (DC) phenotype evidenced by secretion of interleukin-12 from DCs upon activation with tumor antigens from Mito-DCA treated cancer cells. Targeting mitochondrial metabolic inhibitors to the mitochondria could lead to induction of an efficient antitumor immune response, thus introducing the concept of combining glycolysis inhibition with immune system to destroy tumor.

  4. Mitochondria-Targeted Vitamin E Protects Skin from UVB-Irradiation.

    PubMed

    Kim, Won-Serk; Kim, Ikyon; Kim, Wang-Kyun; Choi, Ju-Yeon; Kim, Doo Yeong; Moon, Sung-Guk; Min, Hyung-Keun; Song, Min-Kyu; Sung, Jong-Hyuk

    2016-05-01

    Mitochondria-targeted vitamin E (MVE) is designed to accumulate within mitochondria and is applied to decrease mitochondrial oxidative damage. However, the protective effects of MVE in skin cells have not been identified. We investigated the protective effect of MVE against UVB in dermal fibroblasts and immortalized human keratinocyte cell line (HaCaT). In addition, we studied the wound-healing effect of MVE in animal models. We found that MVE increased the proliferation and survival of fibroblasts at low concentration (i.e., nM ranges). In addition, MVE increased collagen production and downregulated matrix metalloproteinase1. MVE also increased the proliferation and survival of HaCaT cells. UVB increased reactive oxygen species (ROS) production in fibroblasts and HaCaT cells, while MVE decreased ROS production at low concentration. In an animal experiment, MVE accelerated wound healing from laser-induced skin damage. These results collectively suggest that low dose MVE protects skin from UVB irradiation. Therefore, MVE can be developed as a cosmetic raw material.

  5. Mitochondria-targeted antioxidant preserves contractile properties and mitochondrial function of skeletal muscle in aged rats

    PubMed Central

    Javadov, Sabzali; Jang, Sehwan; Rodriguez-Reyes, Natividad; Rodriguez-Zayas, Ana E.; Hernandez, Jessica Soto; Krainz, Tanja; Wipf, Peter; Frontera, Walter

    2015-01-01

    Mitochondrial dysfunction plays a central role in the pathogenesis of sarcopenia associated with a loss of mass and activity of skeletal muscle. In addition to energy deprivation, increased mitochondrial ROS damage proteins and lipids in aged skeletal muscle. Therefore, prevention of mitochondrial ROS is important for potential therapeutic strategies to delay sarcopenia. This study elucidates the pharmacological efficiency of the new developed mitochondria-targeted ROS and electron scavenger, XJB-5-131 (XJB) to restore muscle contractility and mitochondrial function in aged skeletal muscle. Male adult (5-month old) and aged (29-month old) Fischer Brown Norway (F344/BN) rats were treated with XJB for four weeks and contractile properties of single skeletal muscle fibres and activity of mitochondrial ETC complexes were determined at the end of the treatment period. XJB-treated old rats showed higher muscle contractility associated with prevention of protein oxidation in both muscle homogenate and mitochondria compared with untreated counterparts. XJB-treated animals demonstrated a high activity of the respiratory complexes I, III, and IV with no changes in citrate synthase activity. These data demonstrate that mitochondrial ROS play a causal role in muscle weakness, and that a ROS scavenger specifically targeted to mitochondria can reverse age-related alterations of mitochondrial function and improve contractile properties in skeletal muscle. PMID:26415224

  6. Mitochondrial nitric oxide synthase, mitochondrial brain dysfunction in aging, and mitochondria-targeted antioxidants.

    PubMed

    Navarro, Ana; Boveris, Alberto

    2008-01-01

    This paper reviews the current ideas on nitric oxide (NO) physiology in brain and other mammalian organs and on the subcellular distribution of nitric oxide synthases (NOS) emphasizing on the evidence of a mitochondrial NOS isoform (mtNOS) that exhibits a mean activity of 0.86+/-0.09 nmol NO/min x mg protein in 13 mouse and rat organs. Mammalian brain aging is associated with mitochondrial dysfunction, determined as decreased electron transfer and enzymatic activities and as an increased content of phospholipid oxidation products and of protein oxidation/nitration products. Brain mtNOS is the most decreased enzymatic activity upon aging; decreased levels of NO are interpreted as the cause of decreased mitochondrial biogenesis in aged brain. The beneficial effect of high doses of vitamin E on mice survival and neurological function are related to its effect as antioxidant in brain mitochondria and to the preservation of mtNOS activity. Mitochondria-targeted antioxidants, phosphonium cation derivatives and antioxidant tetrapeptides, are reviewed in terms of structures and biological effects.

  7. Mitochondria-Targeted Vitamin E Protects Skin from UVB-Irradiation

    PubMed Central

    Kim, Won-Serk; Kim, Ikyon; Kim, Wang-Kyun; Choi, Ju-Yeon; Kim, Doo Yeong; Moon, Sung-Guk; Min, Hyung-Keun; Song, Min-Kyu; Sung, Jong-Hyuk

    2016-01-01

    Mitochondria-targeted vitamin E (MVE) is designed to accumulate within mitochondria and is applied to decrease mitochondrial oxidative damage. However, the protective effects of MVE in skin cells have not been identified. We investigated the protective effect of MVE against UVB in dermal fibroblasts and immortalized human keratinocyte cell line (HaCaT). In addition, we studied the wound-healing effect of MVE in animal models. We found that MVE increased the proliferation and survival of fibroblasts at low concentration (i.e., nM ranges). In addition, MVE increased collagen production and downregulated matrix metalloproteinase1. MVE also increased the proliferation and survival of HaCaT cells. UVB increased reactive oxygen species (ROS) production in fibroblasts and HaCaT cells, while MVE decreased ROS production at low concentration. In an animal experiment, MVE accelerated wound healing from laser-induced skin damage. These results collectively suggest that low dose MVE protects skin from UVB irradiation. Therefore, MVE can be developed as a cosmetic raw material. PMID:26869457

  8. Mitochondria-targeting photoacoustic therapy using single-walled carbon nanotubes.

    PubMed

    Zhou, Feifan; Wu, Shengnan; Yuan, Yi; Chen, Wei R; Xing, Da

    2012-05-21

    In vitro photoacoustic therapy using modified single-walled carbon nanotubes (SWNTs) as "bomb" agents is a newly reported approach for cancer. Herein, a mitochondria-targeting photoacoustic modality using unmodified SWNTs and its in vitro and in vivo antitumor effect are reported. Unmodified SWNTs can be taken up into cancer cells due to a higher mitochondrial transmembrane potential in cancerous cells than normal cells. Under the irradiation of a 1064 nm pulse laser, 79.4% of cancer cells with intracellular SWNTs die within 20 s, while 82.3% of normal cells without SWNTs remain alive. This modality kills cancer cells mainly by triggering cell apoptosis that initiates from mitochondrial damage, through the depolarization of mitochondria and the subsequent release of cytochrome c after photoacoustic therapy. It is very effective in suppressing tumor growth by selectively destroying tumor tissue without causing epidermis injury. Taken together, these discoveries provide a new method using mitochondria-localized SWNTs as photoacoustic transducers for cancer treatment.

  9. Mitochondria-Targeted Antioxidants and Uncouplers of Oxidative Phosphorylation in Treatment of the Systemic Inflammatory Response Syndrome (SIRS).

    PubMed

    Zakharova, Vlada V; Pletjushkina, Olga Yu; Zinovkin, Roman A; Popova, Ekaterina N; Chernyak, Boris V

    2017-05-01

    Systemic inflammatory response syndrome (SIRS) development is accompanied by mitochondrial dysfunction and excessive ROS production. Mitochondrial dysfunctions also occur in many SIRS-related diseases and may be critical for their pathogenesis; therefore, a use of mitochondria-targeted drugs is a promising trend in SIRS research and therapy. Here, we review recent studies concerning the application of the mitochondria-targeted antioxidants and uncouplers of oxidative phosphorylation in animal models of SIRS and related diseases. We propose that a new class of uncouplers of oxidative phosphorylation, lipophilic cations could be a base for a new generation of drugs for SIRS treatment. J. Cell. Physiol. 232: 904-912, 2017. © 2016 Wiley Periodicals, Inc.

  10. Reduction of early reperfusion injury with the mitochondria-targeting peptide bendavia.

    PubMed

    Brown, David A; Hale, Sharon L; Baines, Christopher P; del Rio, Carlos L; Hamlin, Robert L; Yueyama, Yukie; Kijtawornrat, Anusak; Yeh, Steve T; Frasier, Chad R; Stewart, Luke M; Moukdar, Fatiha; Shaikh, Saame Raza; Fisher-Wellman, Kelsey H; Neufer, P Darrell; Kloner, Robert A

    2014-01-01

    We recently showed that Bendavia, a novel mitochondria-targeting peptide, reduced infarction and no-reflow across several experimental models. The purpose of this study was to determine the therapeutic timing and mechanism of action that underlie Bendavia's cytoprotective property. In rabbits exposed to in vivo ischemia/reperfusion (30/180 min), Bendavia administered 20 minutes prior to reperfusion (0.05 mg/kg/h, intravenously) reduced myocardial infarct size by ∼50% when administered for either 1 or 3 hours of reperfusion. However, when Bendavia perfusion began just 10 minutes after the onset of reperfusion, the protection against infarction and no-reflow was completely lost, indicating that the mechanism of protection is occurring early in reperfusion. Experiments in isolated mouse liver mitochondria found no discernible effect of Bendavia on blocking the permeability transition pore, and studies in isolated heart mitochondria showed no effect of Bendavia on respiratory rates. As Bendavia significantly lowered reactive oxygen species (ROS) levels in isolated heart mitochondria, the ROS-scavenging capacity of Bendavia was compared to well-known ROS scavengers using in vitro (cell-free) systems that enzymatically generate ROS. Across doses ranging from 1 nmol/L to 1 mmol/L, Bendavia showed no discernible ROS-scavenging properties, clearly differentiating itself from prototypical scavengers. In conclusion, Bendavia is a promising candidate to reduce cardiac injury when present at the onset of reperfusion but not after reperfusion has already commenced. Given that both infarction and no-reflow are related to increased cellular ROS, Bendavia's protective mechanism of action likely involves reduced ROS generation (as opposed to augmented scavenging) by endothelial and myocyte mitochondria.

  11. Reduction of early reperfusion injury with the mitochondria-targeting peptide Bendavia

    PubMed Central

    Brown, David A.; Hale, Sharon L.; Baines, Christopher P.; del Rio, Carlos L.; Hamlin, Robert L.; Yueyama, Yukie; Kijtawornrat, Anusak; Yeh, Steve T.; Frasier, Chad R.; Stewart, Luke M.; Moukdar, Fatiha; Shaikh, Saame Raza; Fisher-Wellman, Kelsey H.; Neufer, P. Darrell; Kloner, Robert A.

    2014-01-01

    We recently showed that Bendavia, a novel mitochondria-targeting peptide, reduced infarction and no-reflow across several experimental models. The purpose of this study was to determine the therapeutic timing and mechanism of action that underlie Bendavia’s cytoprotective property. In rabbits exposed to in vivo ischemia/reperfusion (30/180 min), Bendavia administered 20 min prior to reperfusion (0.05mg/kg/hr, i.v.) reduced myocardial infarct size by ~50% when administered for either 1 or 3 hours of reperfusion. However, when Bendavia perfusion began just 10 min after the onset of reperfusion, the protection against infarction and no–reflow was completely lost, indicating that the mechanism of protection is occurring early in reperfusion. Experiments in isolated mouse liver mitochondria found no discernible effect of Bendavia on blocking the permeability transition pore, and studies in isolated heart mitochondria showed no effect of Bendavia on respiratory rates. As Bendavia significantly lowered reactive oxygen species (ROS) levels in isolated heart mitochondria, the ROS-scavenging capacity of Bendavia was compared to well-known ROS scavengers using in vitro (cell-free) systems that enzymatically generate ROS. Across doses ranging from 1nM to 1mM, Bendavia showed no discernible ROS-scavenging properties, clearly differentiating itself from prototypical scavengers. In conclusion, Bendavia is a promising candidate to reduce cardiac injury when present at onset of reperfusion, but not after reperfusion has already commenced. Given that both infarction and no-reflow are related to increased cellular ROS, Bendavia’s protective mechanism of action likely involves reduced ROS generation (as opposed to augmented scavenging) by endothelial and myocyte mitochondria. PMID:24288396

  12. Mitochondria-targeted small molecule SS31: a potential candidate for the treatment of Alzheimer's disease.

    PubMed

    Reddy, P Hemachandra; Manczak, Maria; Kandimalla, Ramesh

    2017-04-15

    The objective of our study was to better understand the protective effects of the mitochondria-targeted tetra-peptide SS31 against amyloid beta (Aβ)-induced mitochondrial and synaptic toxicities in Alzheimer's disease (AD) progression. Using intraperitoneal injections, we administered SS31 to an AD mouse model (APP) over a period of 6 weeks, beginning when the APP mice were 12 months of age. We studied their cortical tissues after SS31 treatment and determined that SS31 crosses the blood brain barrier and reaches mitochondrial sites of free radical production. We also determined: (1) plasma and brain levels of SS31, (2) mRNA levels and levels of mitochondrial dynamics, biogenesis proteins and synaptic proteins, (3) soluble Aβ levels and immunoreactivity of mutant APP and Aβ levels and (4) mitochondrial function by measuring H2O2, lipid peroxidation, cytochrome c oxidase activity and mitochondrial ATP. We found reduced mRNA expression and reduced protein levels of fission genes, and increased levels of mitochondrial fusion, biogenesis and synaptic genes in SS31-treated APP mice relative to SS31-untreated APP mice. Immunofluorescence analysis revealed reduced full-length mutant APP and soluble/insoluble Aβ levels in the SS31-treated APP mice. Sandwich ELISA assays revealed significantly reduced soluble Aβ levels in the SS31-treated APP mice relative to the untreated APP mice. Mitochondrial function was maintained in the SS31-treated APP mice over the 6 weeks of SS31 treatment compared with mitochondrial function in the untreated APP mice. Our findings indicate that SS31 treatment reduces Aβ production, reduces mitochondrial dysfunction, maintains mitochondrial dynamics and enhances mitochondrial biogenesis and synaptic activity in APP mice; and that SS31 may confer protective effects against mitochondrial and synaptic toxicities in APP transgenic mice. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email

  13. The energy blocker inside the power house: Mitochondria targeted delivery of 3-bromopyruvate.

    PubMed

    Marrache, Sean; Dhar, Shanta

    2015-03-01

    A key hallmark of many aggressive cancers is accelerated glucose metabolism. The enzymes that catalyze the first step of glucose metabolism are hexokinases. High levels of hexokinase 2 (HK2) are found in cancer cells, but only in a limited number of normal tissues. Metabolic reprogramming of cancer cells using the energy blocker, 3-bromopyruvate (3-BP) that inhibits HK2 has the potential to provide tumor-specific anticancer agents. However, the unique structural and functional characteristics of mitochondria prohibit selective subcellular targeting of 3-BP to modulate the function of this organelle for therapeutic gain. A mitochondria targeted gold nanoparticle (T-3-BP-AuNP) decorated with 3-BP and delocalized lipophilic triphenylphosphonium cations to target the mitochondrial membrane potential (Δψm) was developed for delivery of 3-BP to cancer cell mitochondria by taking advantage of higher Δψm in cancer cells compared to normal cells. In vitro studies demonstrated enhanced anticancer activity of T-3-BP-AuNPs compared to the non-targeted construct NT-3-BP-AuNP or free 3-BP. The anticancer activity of T-3-BP-AuNP was further enhanced upon laser irradiation by exciting the surface plasmon resonance band of AuNP and thereby utilizing a combination of 3-BP chemotherapeutic and AuNP photothermal effects. The less toxic behavior of T-3-BPNPs in normal mesenchymal stem cells indicated that these NPs preferentially kill cancer cells. T-3-BP-AuNPs showed enhanced ability to modulate cancer cell metabolism by inhibiting glycolysis as well as demolishing mitochondrial oxidative phosphorylation. Our findings demonstrated that concerted chemo-photothermal treatment of glycolytic cancer cells with a single NP capable of targeting mitochondria mediating simultaneous release of a glycolytic inhibitor and photothermal ablation may have promise as a new anticancer therapy.

  14. Reversal of doxorubicin resistance in breast cancer by mitochondria-targeted pH-responsive micelles.

    PubMed

    Yu, Pengcheng; Yu, Haijun; Guo, Chengyue; Cui, Zhirui; Chen, Xianzhi; Yin, Qi; Zhang, Pengcheng; Yang, Xiangliang; Cui, Honggang; Li, Yaping

    2015-03-01

    Chemotherapy is an important approach for clinical cancer treatment. However, the success of chemotherapy is usually hindered by the occurrence of intrinsic or acquired multidrug resistance of cancer cells. Herein, we reported an effective approach to overcome doxorubicin (DOX) resistance in MCF-7/ADR breast cancer using DOX-loaded pH-responsive micelles. The micelles were prepared from a pH-responsive diblock copolymer, poly(ethylene glycol)-block-poly(2-(diisopropylamino)ethyl methacrylate) (PEG-b-PDPA), and a vitamin E derivate (D-α-tocopheryl polyethylene glycol 1000 succinate, TPGS) (denoted as PDPA/TPGS micelles). At neutral pH of 7.4, DOX was loaded into the hydrophobic core of PDPA/TPGS micelles via a film sonication method. After cellular uptake, the DOX payload was released in early endosomes by acidic pH-triggered micelle dissociation. Meanwhile, the TPGS component synergistically improved the cytotoxicity of DOX by targeting mitochondrial organelles and reducing the mitochondrial transmembrane potential. In vitro cell culture experiments using DOX-resistant MCF-7/ADR cells demonstrated that PDPA/TPGS micelles reduced the IC50 of DOX by a sixfold magnitude. In vivo animal studies showed that DOX-loaded PDPA/TPGS micelles (PDPA/TPGS@DOX) inhibited tumor growth more efficiently than free DOX in a nude mouse model bearing orthotopic MCF-7/ADR tumor. All these results imply that the mitochondria-targeted pH-responsive PDPA/TPGS micelles have significant potential for efficiently combating DOX resistance in breast cancer cells. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  15. Mitochondria-targeted antioxidants protect against mechanical ventilation-induced diaphragm weakness.

    PubMed

    Powers, Scott K; Hudson, Matthew B; Nelson, W Bradley; Talbert, Erin E; Min, Kisuk; Szeto, Hazel H; Kavazis, Andreas N; Smuder, Ashley J

    2011-07-01

    Mechanical ventilation is a life-saving intervention used to provide adequate pulmonary ventilation in patients suffering from respiratory failure. However, prolonged mechanical ventilation is associated with significant diaphragmatic weakness resulting from both myofiber atrophy and contractile dysfunction. Although several signaling pathways contribute to diaphragm weakness during mechanical ventilation, it is established that oxidative stress is required for diaphragmatic weakness to occur. Therefore, identifying the site(s) of mechanical ventilation- induced reactive oxygen species production in the diaphragm is important. These experiments tested the hypothesis that elevated mitochondrial reactive oxygen species emission is required for mechanical ventilation-induced oxidative stress, atrophy, and contractile dysfunction in the diaphragm. Cause and effect was determined by preventing mechanical ventilation-induced mitochondrial reactive oxygen species emission in the diaphragm of rats using a novel mitochondria-targeted antioxidant (SS-31). None. Compared to mechanically ventilated animals treated with saline, animals treated with SS-31 were protected against mechanical ventilation-induced mitochondrial dysfunction, oxidative stress, and protease activation in the diaphragm. Importantly, treatment of animals with the mitochondrial antioxidant also protected the diaphragm against mechanical ventilation-induced myofiber atrophy and contractile dysfunction. These results reveal that prevention of mechanical ventilation-induced increases in diaphragmatic mitochondrial reactive oxygen species emission protects the diaphragm from mechanical ventilation-induced diaphragmatic weakness. This important new finding indicates that mitochondria are a primary source of reactive oxygen species production in the diaphragm during prolonged mechanical ventilation. These results could lead to the development of a therapeutic intervention to impede mechanical ventilation

  16. Toxicity of neurons treated with herbicides and neuroprotection by mitochondria-targeted antioxidant SS31.

    PubMed

    Reddy, Tejaswini P; Manczak, Maria; Calkins, Marcus J; Mao, Peizhong; Reddy, Arubala P; Shirendeb, Ulziibat; Park, Byung; Reddy, P Hemachandra

    2011-01-01

    The purpose of this study was to determine the neurotoxicity of two commonly used herbicides: picloram and triclopyr and the neuroprotective effects of the mitochondria-targeted antioxidant, SS31. Using mouse neuroblastoma (N2a) cells and primary neurons from C57BL/6 mice, we investigated the toxicity of these herbicides, and protective effects of SS1 peptide against picloram and triclopyr toxicity. We measured total RNA content, cell viability and mRNA expression of peroxiredoxins, neuroprotective genes, mitochondrial-encoded electron transport chain (ETC) genes in N2a cells treated with herbicides and SS31. Using primary neurons from C57BL/6 mice, neuronal survival was studied in neurons treated with herbicides, in neurons pretreated with SS31 plus treated with herbicides, neurons treated with SS31 alone, and untreated neurons. Significantly decreased total RNA content, and cell viability in N2a cells treated with picloram and triclopyr were found compared to untreated N2a cells. Decreased mRNA expression of neuroprotective genes, and ETC genes in cells treated with herbicides was found compared to untreated cells. Decreased mRNA expression of peroxiredoxins 1-6 in N2a cells treated with picloram was found, suggesting that picloram affects the antioxidant enzymes in N2a cells. Immunofluorescence analysis of primary neurons revealed that decreased neuronal branching and degenerating neurons in neurons treated with picloram and triclopyr. However, neurons pretreated with SS31 prevented degenerative process caused by herbicides. Based on these results, we propose that herbicides--picloram and triclopyr appear to damage neurons, and the SS31 peptide appears to protect neurons from herbicide toxicity.

  17. Toxicity of Neurons Treated with Herbicides and Neuroprotection by Mitochondria-Targeted Antioxidant SS31

    PubMed Central

    Reddy, Tejaswini P.; Manczak, Maria; Calkins, Marcus J.; Mao, Peizhong; Reddy, Arubala P.; Shirendeb, Ulziibat; Park, Byung; Reddy, P. Hemachandra

    2011-01-01

    The purpose of this study was to determine the neurotoxicity of two commonly used herbicides: picloram and triclopyr and the neuroprotective effects of the mitochondria-targeted antioxidant, SS31. Using mouse neuroblastoma (N2a) cells and primary neurons from C57BL/6 mice, we investigated the toxicity of these herbicides, and protective effects of SS1 peptide against picloram and triclopyr toxicity. We measured total RNA content, cell viability and mRNA expression of peroxiredoxins, neuroprotective genes, mitochondrial-encoded electron transport chain (ETC) genes in N2a cells treated with herbicides and SS31. Using primary neurons from C57BL/6 mice, neuronal survival was studied in neurons treated with herbicides, in neurons pretreated with SS31 plus treated with herbicides, neurons treated with SS31 alone, and untreated neurons. Significantly decreased total RNA content, and cell viability in N2a cells treated with picloram and triclopyr were found compared to untreated N2a cells. Decreased mRNA expression of neuroprotective genes, and ETC genes in cells treated with herbicides was found compared to untreated cells. Decreased mRNA expression of peroxiredoxins 1–6 in N2a cells treated with picloram was found, suggesting that picloram affects the antioxidant enzymes in N2a cells. Immunofluorescence analysis of primary neurons revealed that decreased neuronal branching and degenerating neurons in neurons treated with picloram and triclopyr. However, neurons pretreated with SS31 prevented degenerative process caused by herbicides. Based on these results, we propose that herbicides—picloram and triclopyr appear to damage neurons, and the SS31 peptide appears to protect neurons from herbicide toxicity. PMID:21318024

  18. A high-resolution mitochondria-targeting ratiometric fluorescent probe for detection of the endogenous hypochlorous acid

    NASA Astrophysics Data System (ADS)

    Zhou, Liyi; Lu, Dan-Qing; Wang, Qianqian; Hu, Shunqin; Wang, Haifei; Sun, Hongyan; Zhang, Xiaobing

    2016-09-01

    Hypochlorite anion, one of the biologically important reactive oxygen species, plays an essential role in diverse normal biochemical functions and abnormal pathological processes. Herein, an efficient high-resolution mitochondria-targeting ratiometric fluorescent probe for hypochlorous acid detection has been designed, synthesized and characterized. It is easily synthesized by the condensation reaction (Cdbnd C) of a 2-(2-hydroxyphenyl) quinazolin-4(3H)-one fluorophore and a cyanine group (mitochondria-targeting), which made the whole molecular a large Stokes shift (210 nm) and the two well-resolved emission peaks separated by 140 nm. As a result, it is considered as a good candidate for high resolution hypochlorous acid imaging in live cells. The ratiometric fluorescent probe exhibited outstanding features of high sensitivity, high selectivity, rapid response time (within 50 s), and excellent mitochondria-targeting ability. Moreover, the probe can also be successfully applied to imaging endogenously hypochlorous acid in the mitochondria of living cells with low cytotoxicity, and high resolution.

  19. Nitroxide delivery system for Nrf2 activation and skin protection.

    PubMed

    Ben Yehuda Greenwald, Maya; Frušić-Zlotkin, Marina; Soroka, Yoram; Sasson, Shmuel Ben; Bianco-Peled, Havazelet; Bitton, Ronit; Kohen, Ron

    2015-08-01

    Cyclic nitroxides are a large group of compounds composed of diverse stable radicals also known as synthetic antioxidants. Although nitroxides are valuable for use in several skin conditions, in in vivo conditions they have several drawbacks, such as nonspecific dispersion in normal tissue, preferential renal clearance and rapid reduction of the nitroxide to the corresponding hydroxylamine. However, these drawbacks can be easily addressed by encapsulating the nitroxides within microemulsions. This approach would allow nitroxide activity and therefore their valuable effects (e.g. activation of the Keap1-Nrf2-EpRE pathway) to continue. In this work, nitroxides were encapsulated in a microemulsion composed of biocompatible ingredients. The nanometric size and shape of the vehicle microemulsion and nitroxide microemulsion displayed high similarity, indicating that the stability of the microemulsions was preserved. Our studies demonstrated that nitroxide microemulsions were more potent inducers of the Keap1-Nrf2-EpRE pathway than the free nitroxides, causing the activation of phase II enzymes. Moreover, microemulsions containing nitroxides significantly reduced UVB-induced cytotoxicity in the skin. Understanding the mechanism of this improved activity may expand the usage of many other Nrf2 modulating molecules in encapsulated form, as a skin protection strategy against oxidative stress-related conditions.

  20. Nitroxides protect against peroxynitrite-induced nitration and oxidation.

    PubMed

    Sadowska-Bartosz, Izabela; Gajewska, Agnieszka; Skolimowski, Janusz; Szewczyk, Rafał; Bartosz, Grzegorz

    2015-12-01

    Nitroxides are promising compounds for prevention of undesired protein modifications. The aim of this study was to compare the efficiency of 11 nitroxides, derivatives of 2,2,6,6-tetramethylpiperidine-1-oxide (TEMPO) and 2,2,5,5-tetramethylpirrolidine-1-oxyl (PROXYL) in prevention of nitration and oxidation of model compounds and human serum albumin (HSA). Most nitroxides were very efficient in preventing loss of fluorescein fluorescence induced by peroxynitrite (PN) (IC50 in the nanomolar range) and preventing HSA nitration. The loss of fluorescein fluorescence was demonstrated to be due to nitration. Nitroxides were more effective in prevention nitration than oxidation reactions. They showed a concentration window for preventing dihydrorhodamine (DHR) 123 oxidation but exerted a prooxidant effect at both high and low concentrations. No prooxidant effect of nitroxides was seen in prevention of DHR123 oxidation induced by SIN-1. In all essays hydrophobic nitroxides (especially 4-nonylamido-TEMPO and 3-carbamolyl-dehydroPROXYL) showed the lowest efficiency. An exception was the prevention of thiol group oxidation by PN and SIN-1 where hydrophobic nitroxides were the most effective, apparently due to binding to the protein. Nitroxides showed low toxicity to MCF-7 cells. Most nitroxides, except for the most hydrophobic ones, protected cells from the cytotoxic action of SIN-1 and SIN-1-induced protein nitration. These results point to potential usefulness of nitroxides for prevention of PN-induced oxidation and, especially, nitration.

  1. Mitochondria-targeted antioxidants protect against Abeta toxicity in Alzheimer’s disease neurons

    PubMed Central

    Manczak, Maria; Mao, Peizhong; Calkins, Marcus; Cornea, Anda; Arubala, Reddy P.; Murphy, Michael P.; Szeto, Hazel H; Park, Byung; Reddy, P. Hemachandra

    2011-01-01

    Alzheimer’s disease (AD) is a progressive, neurodegenerative disease characterized by progressive decline of memory and cognitive functions. Despite tremendous progress that has been made in understanding disease progression and therapeutics of AD, we still do not have drugs that are capable of slowing its progression. The purpose of our study was to investigate the effects of the mitochondria-targeted antioxidants (MTAs) MitoQ and SS31, and the anti-aging agent resveratrol on neurons from a mouse model of Alzheimer’s disease (AD) (Tg2576 line) and on mouse neuroblastoma (N2a) cells incubated with the amyloid beta (Aβ) peptide. Using electron and confocal microscopy, gene expression analysis, and biochemical methods, we studied mitochondrial structure and function, and neurite outgrowth in N2a cells treated with MitoQ, SS31, and resveratrol, and then incubated with Aβ. In N2a cells only incubated with the Aβ, we found increased expressions of mitochondrial fission genes and decreased expression of fusion genes, and also decreased expression of peroxiredoxins, endogenous cytoprotective antioxidant enzymes. Electron microscopy of the N2a cells incubated with Aβ revealed a significantly increased number of mitochondria, indicating that Aβ fragments mitochondria. Biochemical analysis revealed that function is defective in mitochondria. Neurite outgrowth was significantly decreased in Aβ-incubated N2a cells, indicating that Aβ affects neurite outgrowth. However, in N2a cells treated with MitoQ, SS31, and resveratrol, and then incubated with Aβ, abnormal expression of peroxiredoxins and mitochondrial structural genes were prevented and mitochondrial function was normal; intact mitochondria were present and neurite outgrowth was significantly increased. In primary neurons from amyloid beta precursor protein (AβPP) transgenic mice that were treated with MitoQ and SS31, neurite outgrowth was significantly increased and cyclophilin D expression was significantly

  2. Rapid-scan EPR of immobilized nitroxides.

    PubMed

    Yu, Zhelin; Quine, Richard W; Rinard, George A; Tseitlin, Mark; Elajaili, Hanan; Kathirvelu, Velavan; Clouston, Laura J; Boratyński, Przemysław J; Rajca, Andrzej; Stein, Richard; Mchaourab, Hassane; Eaton, Sandra S; Eaton, Gareth R

    2014-10-01

    X-band electron paramagnetic resonance spectra of immobilized nitroxides were obtained by rapid scan at 293 K. Scan widths were 155 G with 13.4 kHz scan frequency for (14)N-perdeuterated tempone and for T4 lysozyme doubly spin labeled with an iodoacetamide spirocyclohexyl nitroxide and 100 G with 20.9 kHz scan frequency for (15)N-perdeuterated tempone. These wide scans were made possible by modifications to our rapid-scan driver, scan coils made of Litz wire, and the placement of highly conducting aluminum plates on the poles of a Bruker 10″ magnet to reduce resistive losses in the magnet pole faces. For the same data acquisition time, the signal-to-noise for the rapid-scan absorption spectra was about an order of magnitude higher than for continuous wave first-derivative spectra recorded with modulation amplitudes that do not broaden the lineshapes.

  3. Rapid-Scan EPR of Immobilized Nitroxides

    PubMed Central

    Yu, Zhelin; Quine, Richard W.; Rinard, George A.; Tseitlin, Mark; Elajaili, Hanan; Kathirvelu, Velavan; Clouston, Laura J.; Boratyński, Przemysław J.; Rajca, Andrzej; Stein, Richard; Mchaourab, Hassane; Eaton, Sandra S.; Eaton, Gareth R.

    2014-01-01

    X-band electron paramagnetic resonance spectra of immobilized nitroxides were obtained by rapid scan at 293 K. Scan widths were 155 G with 13.4 kHz scan frequency for 14N-perdeuterated tempone and for T4 lysozyme doubly spin labeled with an iodoacetamide spirocyclohexyl nitroxide and 100 G with 20.9 kHz scan frequency for 15N-perdeuterated tempone. These wide scans were made possible by modifications to our rapid-scan driver, scan coils made of Litz wire, and the placement of highly conducting aluminum plates on the poles of a Bruker 10" magnet to reduce resistive losses in the magnet pole faces. For the same data acquisition time, the signal-to-noise for the rapid-scan absorption spectra was about an order of magnitude higher than for continuous wave first-derivative spectra recorded with modulation amplitudes that do not broaden the lineshapes. PMID:25240151

  4. Mitochondria-targeted antioxidant mitotempo protects mitochondrial function against amyloid beta toxicity in primary cultured mouse neurons.

    PubMed

    Hu, Hongtao; Li, Mo

    2016-09-09

    Mitochondrial defects including excess reactive oxygen species (ROS) production and compromised ATP generation are featured pathology in Alzheimer's disease (AD). Amyloid beta (Aβ)-mediated mitochondrial ROS overproduction disrupts intra-neuronal Redox balance, in turn exacerbating mitochondrial dysfunction leading to neuronal injury. Previous studies have found the beneficial effects of mitochondria-targeted antioxidants in preventing mitochondrial dysfunction and neuronal injury in AD animal and cell models, suggesting that mitochondrial ROS scavengers hold promise for the treatment of this neurological disorder. In this study, we have determined that mitotempo, a novel mitochondria-targeted antioxidant protects mitochondrial function from the toxicity of Aβ in primary cultured neurons. Our results showed that Aβ-promoted mitochondrial superoxide production and neuronal lipid oxidation were significantly suppressed by the application of mitotempo. Moreover, mitotempo also demonstrated protective effects on mitochondrial bioenergetics evidenced by preserved mitochondrial membrane potential, cytochrome c oxidase activity as well as ATP production. In addition, the Aβ-induced mitochondrial DNA (mtDNA) depletion and decreased expression levels of mtDNA replication-related DNA polymerase gamma (DNA pol γ) and Twinkle were substantially mitigated by mitotempo. Therefore, our study suggests that elimination of excess mitochondrial ROS rescues mitochondrial function in Aβ-insulted neruons; and mitotempo has the potential to be a promising therapeutic agent to protect mitochondrial and neuronal function in AD. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. [Mitochondria-targeted antioxidant SkQR1 selectively protects MDR-negative cells against ionizing radiation].

    PubMed

    Fetisova, E K; Antoshina, M M; Cherepanynets, V D; Izumov, D S; Kireev, I I; Kireev, R I; Lyamzaev, K G; Riabchenko, N I; Chernyak, B V; Skulachev, V P

    2014-01-01

    Radioprotection appeared to be an important problem of today due to atom energetic development and utilization of radiation material in the industry, science and medicine. It has been shown that mitochondrial targeted antioxidant SkQR1 could attenuate radiation injury of human erythroleukemia K562 cells. Pretreatment with SkQR1 before irradiation decreased DNA double strand breaks formation, diminished the number of chromosomal aberrations and suppressed delayed ROS production. Prevention of oxidative stress and normalization of mitochondrial function by mitochondria-targeted antioxidants may be a potential therapeutic strategy not only against immediate consequences of radiation, but, either against its late consequences such as genomic instability. SkQR1 did not protect against radiation-induced damage the K562 subline with high level of multidrug resistance (MDR) due to SkQR1 extrusion with Pgp 170 MDR pump. We suggest that mitochondria-targeted antioxidants might be used for selective protection of normal cells against radiation-induced damage without interference with radiotherapy of MDR-positive tumors.

  6. Demethyleneberberine, a natural mitochondria-targeted antioxidant, inhibits mitochondrial dysfunction, oxidative stress, and steatosis in alcoholic liver disease mouse model.

    PubMed

    Zhang, Pengcheng; Qiang, Xiaoyan; Zhang, Miao; Ma, Dongshen; Zhao, Zheng; Zhou, Cuisong; Liu, Xie; Li, Ruiyan; Chen, Huan; Zhang, Yubin

    2015-01-01

    Excessive alcohol consumption induces oxidative stress and lipid accumulation in the liver. Mitochondria have long been recognized as the key target for alcoholic liver disease (ALD). Recently, the artificial mitochondria-targeted antioxidant MitoQ has been used to treat ALD effectively in mice. Here, we introduce the natural mitochondria-targeted antioxidant demethyleneberberine (DMB), which has been found in Chinese herb Cortex Phellodendri chinensis. The protective effect of DMB on ALD was evaluated with HepG2 cells and acutely/chronically ethanol-fed mice, mimicking two common patterns of drinking in human. The results showed that DMB, which is composed of a potential antioxidant structure, could penetrate the membrane of mitochondria and accumulate in mitochondria either in vitro or in vivo. Consequently, the acute drinking-caused oxidative stress and mitochondrial dysfunction were significantly ameliorated by DMB. Moreover, we also found that DMB suppressed CYP2E1, hypoxia inducible factor α, and inducible nitric oxide synthase, which contributed to oxidative stress and restored sirtuin 1/AMP-activated protein kinase/peroxisome proliferator-activated receptor-γ coactivator-1α pathway-associated fatty acid oxidation in chronic ethanol-fed mice, which in turn ameliorated lipid peroxidation and macrosteatosis in the liver. Taking these findings together, DMB could serve as a novel and potential therapy for ALD in human beings.

  7. A mitochondria targeting Mn nanoassembly of BODIPY for LDH-A, mitochondria modulated therapy and bimodal imaging of cancer.

    PubMed

    Boison, Daniel; Lu, Wen-Long; Xu, Qin-Mei; Yang, Huang; Huang, Tao; Chen, Qiu-Yun; Gao, Jing; Zhao, Yao

    2016-11-01

    HIF-1α and LDH-A are important targets for hypoxia-driven drug resistance. Mitochondria targeted fluorescent manganese(II)-complexes can be used as potential fluorescence imaging agents, MRI contrast agents and HIF-1α and LDH-A involved anticancer complexes. In this study, a fluorescent manganese(II) nanoparticle, labeled as (PEG-Mn-BDA), was synthesized and used as both fluorescent and MRI imaging agents in cancer cells. In vitro bioassay results indicate that PEG-Mn-BDA was able to inhibit LDH-A activity and depolarize mitochondrial membrane potential with the generation of intracellular ROS, which contributed to the induction of apoptosis. Moreover, the pro-apoptotic protein, caspase 3 was highly expressed. In vivo, PEG-Mn-BDA could also exert inhibition on a mouse hepatocellular carcinoma xenograft. These results suggest that mitochondria targeted PEG-Mn-BDA was able to simultaneously induce selective inhibition on cancer cells and a mouse carcinoma xenograft, label cancer cells with fluorescence and enhance MRI contrast. Therefore, PEG-Mn-BDA is a good candidate for cancer treatment and imaging.

  8. Neuroprotective Effects of Mitochondria-Targeted Plastoquinone and Thymoquinone in a Rat Model of Brain Ischemia/Reperfusion Injury.

    PubMed

    Silachev, Denis N; Plotnikov, Egor Y; Zorova, Ljubava D; Pevzner, Irina B; Sumbatyan, Natalia V; Korshunova, Galina A; Gulyaev, Mikhail V; Pirogov, Yury A; Skulachev, Vladimir P; Zorov, Dmitry B

    2015-08-11

    We explored the neuroprotective properties of natural plant-derived antioxidants plastoquinone and thymoquinone (2-demethylplastoquinone derivative) modified to be specifically accumulated in mitochondria. The modification was performed through chemical conjugation of the quinones with penetrating cations: Rhodamine 19 or tetraphenylphosphonium. Neuroprotective properties were evaluated in a model of middle cerebral artery occlusion. We demonstrate that the mitochondria-targeted compounds, introduced immediately after reperfusion, possess various neuroprotective potencies as judged by the lower brain damage and higher neurological status. Plastoquinone derivatives conjugated with rhodamine were the most efficient, and the least efficiency was shown by antioxidants conjugated with tetraphenylphosphonium. Antioxidants were administered intraperitoneally or intranasally with the latter demonstrating a high level of penetration into the brain tissue. The therapeutic effects of both ways of administration were similar. Long-term administration of antioxidants in low doses reduced the neurological deficit, but had no effect on the volume of brain damage. At present, cationic decylrhodamine derivatives of plastoquinone appear to be the most promising anti-ischemic mitochondria-targeted drugs of the quinone family. We suggest these antioxidants could be potentially used for a stroke treatment.

  9. Nitroxide amide-BODIPY probe behavior in fibroblasts analyzed by advanced fluorescence microscopy.

    PubMed

    Liras, M; Simoncelli, S; Rivas-Aravena, A; García, O; Scaiano, J C; Alarcon, E I; Aspée, A

    2016-04-26

    A novel synthesized nitroxide amide-BODIPY prefluorescent probe was used to study cellular redox balance that modulates nitroxide/hydroxylamine ratio in cultured human fibroblasts. FLIM quantitatively differentiated between nitroxide states of the cytoplasm-localized probe imaged by TIRF, monitoring nitroxide depletion by hydrogen peroxide; eluding incorrect interpretation if only fluorescence intensity is considered.

  10. Identification of functionally important amino acid residues in the mitochondria targeting sequence of Hepatitis B virus X protein

    SciTech Connect

    Li, Sai Kam; Ho, Sai Fan; Tsui, Kwok Wing; Fung, Kwok Pui; Waye, M.Y. Mary

    2008-11-10

    Chronic hepatitis B virus (HBV) infection has been strongly associated with hepatocellular carcinoma (HCC) and the X protein (HBx) is thought to mediate the cellular changes associated with carcinogenesis. Recently, isolation of the hepatitis B virus integrants from HCC tissue by others have established the fact that the X gene is often truncated at its C-terminus. Expression of the GFP fusion proteins of HBx and its truncation mutants with a GFP tag in human liver cell-lines in this study revealed that the C-terminus of HBx is indispensable for its specific localization in the mitochondria. A crucial region of seven amino acids at the C-terminus has been mapped out in which the cysteine residue at position 115 serves as the most important residue for the subcellular localization. When cysteine 115 of HBx is mutated to alanine the mitochondria targeting property of HBx is abrogated.

  11. Mitochondria-targeted cancer therapy using a light-up probe with aggregation-induced-emission characteristics.

    PubMed

    Hu, Qinglian; Gao, Meng; Feng, Guangxue; Liu, Bin

    2014-12-15

    Subcellular organelle-specific reagents for simultaneous tumor targeting, imaging, and treatment are of enormous interest in cancer therapy. Herein, we present a mitochondria-targeting probe (AIE-mito-TPP) by conjugating a triphenylphosphine (TPP) with a fluorogen which can undergo aggregation-induced emission (AIE). Owing to the more negative mitochondrial membrane potential of cancer cells than normal cells, the AIE-mito-TPP probe can selectively accumulate in cancer-cell mitochondria and light up its fluorescence. More importantly, the probe exhibits selective cytotoxicity for studied cancer cells over normal cells. The high potency of AIE-mito-TPP correlates with its strong ability to aggregate in mitochondria, which can efficiently decrease the mitochondria membrane potential and increase the level of intracellular reactive oxygen species (ROS) in cancer cells. The mitochondrial light-up probe provides a unique strategy for potential image-guided therapy of cancer cells.

  12. Design and Synthesis of a Mitochondria-Targeted Mimic of Glutathione Peroxidase, MitoEbselen-2, as a Radiation Mitigator.

    PubMed

    Stoyanovsky, Detcho A; Jiang, Jianfei; Murphy, Michael P; Epperly, Michael; Zhang, Xiaolan; Li, Song; Greenberger, Joel; Kagan, Valerian; Bayır, Hülya

    2014-12-11

    Ionizing radiation (IR) triggers mitochondrial overproduction of H2O2 and accumulation of lipid hydroperoxides leading to the induction of apoptotic and necroptotic cell death pathways. Given the high catalytic efficiency of the seleno-enzyme glutathione peroxidase (Gpx) toward reduction of lipid hydroperoxides and H2O2, we tested the potential of mitochondria-targeted derivatives of ebselen to mitigate the deleterious effects of IR. We report that 2-[[2-[4-(3-oxo-1,2-benzoselenazol-2-yl)phenyl]acetyl]amino]ethyl-triphenyl-phosphonium chloride (MitoPeroxidase 2) was effective in reducing lipid hydroperoxides, preventing apoptotic cell death, and, when administered 24 h postirradiation, increased the survival of mice exposed to whole body γ-irradiation.

  13. The mitochondria-targeted antioxidant MitoQ decreases features of the metabolic syndrome in ATM+/-/ApoE-/- mice.

    PubMed

    Mercer, John R; Yu, Emma; Figg, Nichola; Cheng, Kian-Kai; Prime, Tracy A; Griffin, Julian L; Masoodi, Mojgan; Vidal-Puig, Antonio; Murphy, Michael P; Bennett, Martin R

    2012-03-01

    A number of recent studies suggest that mitochondrial oxidative damage may be associated with atherosclerosis and the metabolic syndrome. However, much of the evidence linking mitochondrial oxidative damage and excess reactive oxygen species (ROS) with these pathologies is circumstantial. Consequently the importance of mitochondrial ROS in the etiology of these disorders is unclear. Furthermore, the potential of decreasing mitochondrial ROS as a therapy for these indications is not known. We assessed the impact of decreasing mitochondrial oxidative damage and ROS with the mitochondria-targeted antioxidant MitoQ in models of atherosclerosis and the metabolic syndrome (fat-fed ApoE(-/-) mice and ATM(+/-)/ApoE(-/-) mice, which are also haploinsufficient for the protein kinase, ataxia telangiectasia mutated (ATM). MitoQ administered orally for 14weeks prevented the increased adiposity, hypercholesterolemia, and hypertriglyceridemia associated with the metabolic syndrome. MitoQ also corrected hyperglycemia and hepatic steatosis, induced changes in multiple metabolically relevant lipid species, and decreased DNA oxidative damage (8-oxo-G) in multiple organs. Although MitoQ did not affect overall atherosclerotic plaque area in fat-fed ATM(+/+)/ApoE(-/-) and ATM(+/-)/ApoE(-/-) mice, MitoQ reduced the macrophage content and cell proliferation within plaques and 8-oxo-G. MitoQ also significantly reduced mtDNA oxidative damage in the liver. Our data suggest that MitoQ inhibits the development of multiple features of the metabolic syndrome in these mice by affecting redox signaling pathways that depend on mitochondrial ROS such as hydrogen peroxide. These findings strengthen the growing view that elevated mitochondrial ROS contributes to the etiology of the metabolic syndrome and suggest a potential therapeutic role for mitochondria-targeted antioxidants. Copyright © 2011 Elsevier Inc. All rights reserved.

  14. Mitochondria-targeted plastoquinone derivative SkQ(1) decreases ischemia-reperfusion injury during liver hypothermic storage for transplantation.

    PubMed

    Cherkashina, D V; Sosimchik, I A; Semenchenko, O A; Volina, V V; Petrenko, A Yu

    2011-09-01

    The ability of the mitochondria-targeted plastoquinone derivative 10-(6'-plastoquinonyl)decyl triphenylphosphonium (SkQ(1)) to decrease ischemia-reperfusion injury in isolated liver during hypothermic storage (HS) was studied. Rat liver was stored for 24 h at 4°C without or in the presence of 1 μM SkQ(1) with following reperfusion for 60 min at 37°C. The presence in the storage medium of SkQ(1) significantly decreased spontaneous production of reactive oxygen species and intensity of lipid peroxidation in the liver during HS and reperfusion. The GSH level after HS in solution with SkQ(1) was reliably higher, but reperfusion leveled this effect. At all stages of experiment the presence of SkQ(1) did not prevent the decrease of antioxidant enzyme activities such as catalase, GSH peroxidase, GSH reductase, and glucose-6-phosphate dehydrogenase. The addition of SkQ(1) to the storage medium improved energetic function of the liver, as was revealed in increased respiratory control index of mitochondria and ATP level. SkQ(1) exhibited positive effect on the liver secretory function and morphology after HS as revealed in enhanced bile flow rate during reperfusion and partial recovery of organ architectonics and state of liver sinusoids and hepatocytes. The data point to promising application of mitochondria-targeted antioxidants for correction of the ischemia-reperfusion injury of isolated liver during long-term cold storage before transplantation.

  15. Plant cell death caused by fungal, bacterial, and viral elicitors: protective effect of mitochondria-targeted quinones.

    PubMed

    Kiselevsky, D B; Frolova, O Yu; Solovyev, A G; Dorokhov, Yu L; Morozov, S Yu; Samuilov, V D

    2014-12-01

    Chitosan (partially deacetylated chitin), a component of fungal cell walls, caused epidermal cell (EC) death in the leaves of pea (Pisum sativum L.) and tobacco Nicotiana tabacum or Nicotiana benthamiana detected by destruction of cell nuclei. The mitochondria-targeted quinone SkQ1 prevented the destruction of EC nuclei induced by chitosan. Chitosan increased and SkQ1 suppressed the activity of protein kinases in N. benthamiana and P. sativum and eliminated the effect of chitosan. Chitosan induced the generation of reactive oxygen species (ROS) in the guard cells (GC) of pea plants. Treatment with chitosan or H2O2 did not cause destruction of GC nuclei; however, it resulted in disruption of the permeability barrier of the plasma membrane detected by propidium iodide fluorescence. Treatment with bacterial lipopolysaccharide but not peptidoglycan caused destruction of pea EC nuclei, which was prevented by SkQ1. Leaves of tobacco plants containing the N gene responsible for resistance to tobacco mosaic virus (TMV) were infiltrated with Agrobacterium tumefaciens cells. These cells contained a genetic construct with the gene of the helicase domain of TMV replicase (p50); its protein product p50 is a target for the N-gene product. As a result, the hypersensitive response (HR) was initiated. The HR manifested itself in the death of leaves and was suppressed by SkQ3. Treatment of tobacco epidermal peels with the A. tumefaciens cells for the p50 gene expression stimulated the destruction of EC nuclei, which was inhibited by SkQ1 or SkQ3. The p50-lacking A. tumefaciens cells did not induce the destruction of EC nuclei. The protective effect of mitochondria-targeted antioxidants SkQ1 and SkQ3 demonstrates the involvement of mitochondria and their ROS in programmed cell death caused by pathogen elicitors.

  16. Inhibition of myeloperoxidase-mediated hypochlorous acid production by nitroxides.

    PubMed

    Rees, Martin D; Bottle, Steven E; Fairfull-Smith, Kathryn E; Malle, Ernst; Whitelock, John M; Davies, Michael J

    2009-06-12

    Tissue damage resulting from the extracellular production of HOCl (hypochlorous acid) by the MPO (myeloperoxidase)-hydrogen peroxide-chloride system of activated phagocytes is implicated as a key event in the progression of a number of human inflammatory diseases. Consequently, there is considerable interest in the development of therapeutically useful MPO inhibitors. Nitroxides are well established antioxidant compounds of low toxicity that can attenuate oxidative damage in animal models of inflammatory disease. They are believed to exert protective effects principally by acting as superoxide dismutase mimetics or radical scavengers. However, we show here that nitroxides can also potently inhibit MPO-mediated HOCl production, with the nitroxide 4-aminoTEMPO inhibiting HOCl production by MPO and by neutrophils with IC50 values of approx. 1 and 6 microM respectively. Structure-activity relationships were determined for a range of aliphatic and aromatic nitroxides, and inhibition of oxidative damage to two biologically-important protein targets (albumin and perlecan) are demonstrated. Inhibition was shown to involve one-electron oxidation of the nitroxides by the compound I form of MPO and accumulation of compound II. Haem destruction was also observed with some nitroxides. Inhibition of neutrophil HOCl production by nitroxides was antagonized by neutrophil-derived superoxide, with this attributed to superoxide-mediated reduction of compound II. This effect was marginal with 4-aminoTEMPO, probably due to the efficient superoxide dismutase-mimetic activity of this nitroxide. Overall, these data indicate that nitroxides have considerable promise as therapeutic agents for the inhibition of MPO-mediated damage in inflammatory diseases.

  17. Inhibition of myeloperoxidase-mediated hypochlorous acid production by nitroxides

    PubMed Central

    Rees, Martin D.; Bottle, Steven E.; Fairfull-Smith, Kathryn E.; Malle, Ernst; Whitelock, John M.; Davies, Michael J.

    2014-01-01

    Tissue damage resulting from the extracellular production of HOCl (hypochlorous acid) by the MPO (myeloperoxidase)-hydrogen peroxide-chloride system of activated phagocytes is implicated as a key event in the progression of a number of human inflammatory diseases. Consequently, there is considerable interest in the development of therapeutically useful MPO inhibitors. Nitroxides are well established antioxidant compounds of low toxicity that can attenuate oxidative damage in animal models of inflammatory disease. They are believed to exert protective effects principally by acting as superoxide dismutase mimetics or radical scavengers. However, we show here that nitroxides can also potently inhibit MPO-mediated HOCl production, with the nitroxide 4-aminoTEMPO inhibiting HOCl production by MPO and by neutrophils with IC50 values of approx. 1 and 6 μM respectively. Structure–activity relationships were determined for a range of aliphatic and aromatic nitroxides, and inhibition of oxidative damage to two biologically-important protein targets (albumin and perlecan) are demonstrated. Inhibition was shown to involve one-electron oxidation of the nitroxides by the compound I form of MPO and accumulation of compound II. Haem destruction was also observed with some nitroxides. Inhibition of neutrophil HOCl production by nitroxides was antagonized by neutrophil-derived superoxide, with this attributed to superoxide-mediated reduction of compound II. This effect was marginal with 4-aminoTEMPO, probably due to the efficient superoxide dismutase-mimetic activity of this nitroxide. Overall, these data indicate that nitroxides have considerable promise as therapeutic agents for the inhibition of MPO-mediated damage in inflammatory diseases. PMID:19379130

  18. Ruthenium nitrosyl functionalized graphene quantum dots as an efficient nanoplatform for NIR-light-controlled and mitochondria-targeted delivery of nitric oxide combined with photothermal therapy.

    PubMed

    Guo, Min; Xiang, Hui-Jing; Wang, Yi; Zhang, Qian-Ling; An, Lu; Yang, Shi-Ping; Ma, Yinchu; Wang, Yucai; Liu, Jin-Gang

    2017-03-18

    A mitochondria-targeting nanoplatform for near-infrared-light-controlled release of nitric oxide accompanied by photothermal therapy was developed, which consists of ruthenium nitrosyl functionalized N-doped graphene quantum dots and a triphenylphosphonium moiety. The nanoplatform demonstrated both in vitro and in vivo anti-tumor efficacy upon irradiation with 808 nm light.

  19. Photoactivatable cytotoxic agents derived from mitochondria-targeting luminescent iridium(III) poly(ethylene glycol) complexes modified with a nitrobenzyl linkage.

    PubMed

    Tso, Karson Ka-Shun; Leung, Kam-Keung; Liu, Hua-Wei; Lo, Kenneth Kam-Wing

    2016-03-25

    Two novel photoactivatable mitochondria-targeting luminescent iridium(III) poly(ethylene glycol) (PEG) complexes incorporated with a nitrobenzyl group were designed. They showed minimal cytotoxic activity in the dark, but became significantly cytotoxic upon irradiation due to the release of the PEG pendants.

  20. Tritiation and Stability Measurements of Nitroxide for Betavoltaic Cells

    DTIC Science & Technology

    2016-09-01

    GaN surrounds tritiated nitroxide on all 6 sides of the tritiated compound cube . .....................................23 Fig. B.1 Percentage of beta...These values will account for 6 faces of the tritiated compound cube . These values will be compared to past and present metal hydrides. Fig...µm. GaN surrounds tritiated nitroxide on all 6 sides of the tritiated compound cube . In the initial phase of this program in which we exceeded all

  1. Cross relaxation in nitroxide spin labels

    NASA Astrophysics Data System (ADS)

    Marsh, Derek

    2016-11-01

    Cross relaxation, and mI -dependence of the intrinsic electron spin-lattice relaxation rate We , are incorporated explicitly into the rate equations for the electron-spin population differences that govern the saturation behaviour of 14N- and 15N-nitroxide spin labels. Both prove important in spin-label EPR and ELDOR, particularly for saturation recovery studies. Neither for saturation recovery, nor for CW-saturation EPR and CW-ELDOR, can cross relaxation be described simply by increasing the value of We , the intrinsic spin-lattice relaxation rate. Independence of the saturation recovery rates from the hyperfine line pumped or observed follows directly from solution of the rate equations including cross relaxation, even when the intrinsic spin-lattice relaxation rate We is mI -dependent.

  2. Mitochondria-targeted antioxidant SkQ1 reduces age-related alterations in the ultrastructure of the lacrimal gland

    PubMed Central

    Bakeeva, Lora E.; Eldarov, Chupalav M.; Vangely, Irina M.; Kolosova, Nataliya G.; Vays, Valeriya B.

    2016-01-01

    Dry eye syndrome is an eye disorder affecting many people at an old age. Because dry eye syndrome is accelerated by aging, a useful approach to the prevention of this syndrome may be an intervention into the aging process. Previously, we showed that the mitochondria-targeted antioxidant SkQ1 delays manifestations of aging and inhibits the development of age-related diseases including dry eye syndrome. Nevertheless, the link between SkQ1's effects and its suppression of age-related changes in the lacrimal gland remains unclear. Here we demonstrated that dietary supplementation with SkQ1 (250 nmol/[kg body weight] daily) starting at age 1.5 months significantly alleviated the pathological changes in lacrimal glands of Wistar rats by age 24 months. By this age, lacrimal glands underwent dramatic deterioration of the ultrastructure that was indicative of irreversible disturbances in these glands' functioning. In contrast, in SkQ1-treated rats, the ultrastructure of the lacrimal gland was similar to that in much younger rats. Morphometric analysis of electron-microscopic specimens of lacrimal glands revealed the presence of numerous secretory granules in acinar cells and a significant increase in the number of operating intercalary ducts. Our results confirm that dietary supplementation with SkQ1 is a promising approach to healthy ageing and to prevention of aberrations in the lacrimal gland that underlie dry eye syndrome. PMID:27852065

  3. Abnormal mitochondrial dynamics and synaptic degeneration as early events in Alzheimer's disease: implications to mitochondria-targeted antioxidant therapeutics.

    PubMed

    Reddy, P Hemachandra; Tripathi, Raghav; Troung, Quang; Tirumala, Karuna; Reddy, Tejaswini P; Anekonda, Vishwanath; Shirendeb, Ulziibat P; Calkins, Marcus J; Reddy, Arubala P; Mao, Peizhong; Manczak, Maria

    2012-05-01

    Synaptic pathology and mitochondrial oxidative damage are early events in Alzheimer's disease (AD) progression. Loss of synapses and synaptic damage are the best correlates of cognitive deficits found in AD patients. Recent research on amyloid beta (Aβ) and mitochondria in AD revealed that Aβ accumulates in synapses and synaptic mitochondria, leading to abnormal mitochondrial dynamics and synaptic degeneration in AD neurons. Further, recent studies using live-cell imaging and primary neurons from amyloid beta precursor protein (AβPP) transgenic mice revealed reduced mitochondrial mass, defective axonal transport of mitochondria and synaptic degeneration, indicating that Aβ is responsible for mitochondrial and synaptic deficiencies. Tremendous progress has been made in studying antioxidant approaches in mouse models of AD and clinical trials of AD patients. This article highlights the recent developments made in Aβ-induced abnormal mitochondrial dynamics, defective mitochondrial biogenesis, impaired axonal transport and synaptic deficiencies in AD. This article also focuses on mitochondrial approaches in treating AD, and also discusses latest research on mitochondria-targeted antioxidants in AD. This article is part of a Special Issue entitled: Antioxidants and Antioxidant Treatment in Disease.

  4. A mitochondria-targeting artemisinin derivative with sharply increased antitumor but depressed anti-yeast and anti-malaria activities

    PubMed Central

    Sun, Chen; Cao, Yu; Zhu, Pan; Zhou, Bing

    2017-01-01

    The potent anti-malarial drug artemisinins are additionally anti-tumorigenic and inhibitory to yeast growth. The action mechanism of artemisinins, however, is not well understood. Heme and mitochondrial membrane are both suggested to be involved in the action of artemisinins. Because heme is also synthesized in the mitochondrion, mitochondria appear to be a critical organelle for artemisinins’ activities. In this study, we synthesized a mitochondria-targeting artemisinin derivative by conjugating triphenylphosphonium (TPP) to artelinic acid (ARTa). ARTa-TPP displays far more potent anti-tumorigenic activity than its parent compound. In contrast, ARTa-TPP is much less active against yeast respiration growth and malarial parasites. Notably, ARTa-TPP is also associated with increased toxicity to other kinds of control mammalian cells. These results suggest divergent action modes for artemisinins against cancer cells and malaria or yeast cells. We conclude that mitochondrial targeting could substantially elevate the anticancer potency of artemisinins, but the accompanied increased toxicity to normal cells raises an alert. The mechanism regarding the opposing effects of TPP conjugation to ARTa on its anticancer and anti-malarial/anti-yeast potencies is discussed based on our current understandings of artemisinins’ action. PMID:28368011

  5. Phosphorescent iridium(III)-bis-N-heterocyclic carbene complexes as mitochondria-targeted theranostic and photodynamic anticancer agents.

    PubMed

    Li, Yi; Tan, Cai-Ping; Zhang, Wei; He, Liang; Ji, Liang-Nian; Mao, Zong-Wan

    2015-01-01

    Mitochondria-targeted compounds represent a promising approach to target tumors selectively and overcome resistance to current anticancer therapies. In this work, three cyclometalated iridium(III) complexes (1-3) containing bis-N-heterocyclic carbene (NHC) ligands have been explored as theranostic and photodynamic agents targeting mitochondria. These complexes display rich photophysical properties, which greatly facilitates the study of their intracellular fate. All three complexes are more cytotoxic than cisplatin against the cancer cells screened. 1-3 can penetrate into human cervical carcinoma (HeLa) cells quickly and efficiently, and they can carry out theranostic functions by simultaneously inducing and monitoring the morphological changes in mitochondria. Mechanism studies show that these complexes exert their anticancer efficacy by initiating a cascade of events related to mitochondrial dysfunction. Additionally, they display up to 3 orders of magnitude higher cytotoxicity upon irradiation at 365 nm, which is so far the highest photocytotoxic responses reported for iridium complexes. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. A mitochondria-targeting artemisinin derivative with sharply increased antitumor but depressed anti-yeast and anti-malaria activities.

    PubMed

    Sun, Chen; Cao, Yu; Zhu, Pan; Zhou, Bing

    2017-04-03

    The potent anti-malarial drug artemisinins are additionally anti-tumorigenic and inhibitory to yeast growth. The action mechanism of artemisinins, however, is not well understood. Heme and mitochondrial membrane are both suggested to be involved in the action of artemisinins. Because heme is also synthesized in the mitochondrion, mitochondria appear to be a critical organelle for artemisinins' activities. In this study, we synthesized a mitochondria-targeting artemisinin derivative by conjugating triphenylphosphonium (TPP) to artelinic acid (ARTa). ARTa-TPP displays far more potent anti-tumorigenic activity than its parent compound. In contrast, ARTa-TPP is much less active against yeast respiration growth and malarial parasites. Notably, ARTa-TPP is also associated with increased toxicity to other kinds of control mammalian cells. These results suggest divergent action modes for artemisinins against cancer cells and malaria or yeast cells. We conclude that mitochondrial targeting could substantially elevate the anticancer potency of artemisinins, but the accompanied increased toxicity to normal cells raises an alert. The mechanism regarding the opposing effects of TPP conjugation to ARTa on its anticancer and anti-malarial/anti-yeast potencies is discussed based on our current understandings of artemisinins' action.

  7. Surface plasmon resonance-induced photoactivation of gold nanoparticles as mitochondria-targeted therapeutic agents for pancreatic cancer.

    PubMed

    Mocan, Lucian; Ilie, Ioana; Tabaran, Flaviu A; Dana, Bartos; Zaharie, Florin; Zdrehus, Claudiu; Puia, Cosmin; Mocan, Teodora; Muntean, Valentin; Teodora, Pop; Ofelia, Mosteanu; Marcel, Tantau; Iancu, Cornel

    2013-12-01

    Noble metal nanoparticles such as gold nanoparticles can strongly absorb light in the visible region by inducing coherent collective oscillation of conduction band electrons in strong resonance with visible frequencies of light. This phenomenon is frequently termed as surface plasmon resonance (SPR). The main objective was to study the effects of laser photoactivated gold nanoparticles (by means of SPR) on human pancreatic cancer cells. Gold nanoparticles obtained using standard wet chemical methods (with sodium borohydride as a reducing agent) underwent photoexcitation using 2w 808 nm laser and further administered to 1.4E7 pancreatic cancer cell lines. Flow cytometry, transmission electron microscopy, phase contrast microscopy, quantitative proteomics and confocal microscopy combined with immunochemical staining were used to examine the interaction between photo excited gold nanoparticles and pancreatic cancer cells. The study shows that phonon-phonon interactions following laser photoexcitation of gold nanoparticles exhibit increased intracellular uptake, as well as mitochondrial swelling, closely followed by mitochondrial inner membrane permeabilization and depolarization. This unique data may represent a major step in mitochondria-targeted anticancer therapies using laser-activated gold nanoparticles.

  8. Nitroxide malonate methanofullerene as biomimetic model of interaction of nitroxide species with antioxidants.

    PubMed

    Melnikova, N B; Korobko, V M; Gulenova, M V; Gubskaya, V P; Fazlleeva, G M; Zhiltsova, O E; Kochetkov, E N; Poddel'sky, A I; Nuretdinov, I A

    2015-12-01

    Bis-nitroxide malonate methanofullerene (NO)2-MF was studied as a biomimetic model of reduction-oxidation activity with natural compounds-cytochrome c (cyt c), dihydroquercetin (DHQ), ascorbic acid (AA) and synthetic drug-1-(β-oxyethyl)-4,6-dimethyl-1,2-dihydro-2-oxopyrimidine (xymedon(®)). (NO)2-MF may be used as the component of Langmuir monolayers on an aqueous subphase and as the adsorbate on silica gel. The activity of (NO)2-MF in the reaction with cyt c was compared with the effect of nitroxide species such as gaseous nitric oxide, 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO) by using UV-vis and EPR-spectra. It has been shown, that iron(III) in cyt c(3+) under action (NO)2-MF was reduced up to iron(II), similar effect was observed under the influence of gaseous NO in aqueous solution, but reduction of iron(III) in heme cyt c was reversible in the presence of TEMPO. Therefore, the state of Fe-heme in cyt c can be used as the indicator of the interaction of cyt c with nitroxide species in vitro. The interaction of cyt c, DHQ, xymedon(®) with (NO)2-MF monolayers was confirmed by the increasing of limiting area А0 from 0.88 nm(2) up to 1.70 nm(2) of (NO)2-MF on the aqueous subphase, by the paramagnetism and UV-vis spectral data changes. These results can be explained by appearance of oxoammonium ion (NO(+))2-MF adlayers and monolayers. The antioxidant and regenerating effects were shown when treating wounds by xymedon(®) in the presence of additives (0.001%) of (NO)2-MF in the experiments on the rats.

  9. pH-Responsive de-PEGylated nanoparticles based on triphenylphosphine-quercetin self-assemblies for mitochondria-targeted cancer therapy.

    PubMed

    Xing, Lei; Lyu, Jin-Yuan; Yang, Yue; Cui, Peng-Fei; Gu, Liu-Qing; Qiao, Jian-Bin; He, Yu-Jing; Zhang, Tian-Qi; Sun, Minjie; Lu, Jin-Jian; Xu, Xiaojun; Liu, Yu; Jiang, Hu-Lin

    2017-08-11

    We have developed mitochondria-targeted self-assembled nanoparticles (NPs) based on amphiphilic triphenylphosphine-quercetin (TPP-Que) conjugates, which were further modified by poly(ethylene glycol) via a pH-responsive coordination bond to form TQ-PEG NPs. And it is revealed that the TQ-PEG NPs were more effective therapeutic agents compared with Que in vitro and in vivo.

  10. Mitochondria-targeted molecules MitoQ and SS31 reduce mutant huntingtin-induced mitochondrial toxicity and synaptic damage in Huntington's disease.

    PubMed

    Yin, Xiangling; Manczak, Maria; Reddy, P Hemachandra

    2016-05-01

    The objective of this study was to determine the protective effects of the mitochondria-targeted molecules MitoQ and SS31 in striatal neurons that stably express mutant huntingtin (Htt) (STHDhQ111/Q111) in Huntington's disease (HD). We studied mitochondrial and synaptic activities by measuring mRNA and the protein levels of mitochondrial and synaptic genes, mitochondrial function, and ultra-structural changes in MitoQ- and SS31-treated mutant Htt neurons relative to untreated mutant Htt neurons. We used gene expression analysis, biochemical methods, transmission electron microscopy (TEM) and confocal microscopy methods. In the MitoQ- and SS31-treated mutant Htt neurons, fission genes Drp1 and Fis1 were down-regulated, and fusion genes Mfn1, Mfn2 and Opa1 were up-regulated relative to untreated neurons, suggesting that mitochondria-targeted molecules reduce fission activity. Interestingly, the mitochondrial biogenesis genes PGC1α, PGC1β, Nrf1, Nrf2 and TFAM were up-regulated in MitoQ- and SS31-treated mutant Htt neurons. The synaptic genes synaptophysin and PSD95 were up-regulated, and mitochondrial function was normal in the MitoQ- and SS31-treated mutant Htt neurons. Immunoblotting findings of mitochondrial and synaptic proteins agreed with the mRNA findings. TEM studies revealed decreased numbers of structurally intact mitochondria in MitoQ- and SS31-treated mutant Htt neurons. These findings suggest that mitochondria-targeted molecules MitoQ and SS31 are protective against mutant Htt-induced mitochondrial and synaptic damage in HD neurons, and these mitochondria-targeted molecules are potential therapeutic molecules for the treatment of HD neurons.

  11. Designing inhibitors of cytochrome c/cardiolipin peroxidase complexes: mitochondria-targeted imidazole-substituted fatty acids.

    PubMed

    Jiang, Jianfei; Bakan, Ahmet; Kapralov, Alexandr A; Silva, K Ishara; Huang, Zhentai; Amoscato, Andrew A; Peterson, James; Garapati, Venkata Krishna; Saxena, Sunil; Bayir, Hülya; Atkinson, Jeffrey; Bahar, Ivet; Kagan, Valerian E

    2014-06-01

    Mitochondria have emerged as the major regulatory platform responsible for the coordination of numerous metabolic reactions as well as cell death processes, whereby the execution of intrinsic apoptosis includes the production of reactive oxygen species fueling oxidation of cardiolipin (CL) catalyzed by cytochrome (Cyt) c. As this oxidation occurs within the peroxidase complex of Cyt c with CL, the latter represents a promising target for the discovery and design of drugs with antiapoptotic mechanisms of action. In this work, we designed and synthesized a new group of mitochondria-targeted imidazole-substituted analogs of stearic acid TPP-n-ISAs with various positions of the attached imidazole group on the fatty acid (n = 6, 8, 10, 13, and 14). By using a combination of absorption spectroscopy and EPR protocols (continuous wave electron paramagnetic resonance and electron spin echo envelope modulation) we demonstrated that TPP-n-ISAs indeed were able to potently suppress CL-induced structural rearrangements in Cyt c, paving the way to its peroxidase competence. TPP-n-ISA analogs preserved the low-spin hexa-coordinated heme-iron state in Cyt c/CL complexes whereby TPP-6-ISA displayed a significantly more effective preservation pattern than TPP-14-ISA. Elucidation of these intermolecular stabilization mechanisms of Cyt c identified TPP-6-ISA as an effective inhibitor of the peroxidase function of Cyt c/CL complexes with a significant antiapoptotic potential realized in mouse embryonic cells exposed to ionizing irradiation. These experimental findings were detailed and supported by all-atom molecular dynamics simulations. Based on the experimental data and computation predictions, we identified TPP-6-ISA as a candidate drug with optimized antiapoptotic potency. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Mitochondria-targeted antioxidant MitoQ ameliorates experimental mouse colitis by suppressing NLRP3 inflammasome-mediated inflammatory cytokines

    PubMed Central

    2013-01-01

    Background MitoQ is a mitochondria-targeted derivative of the antioxidant ubiquinone, with antioxidant and anti-apoptotic functions. Reactive oxygen species are involved in many inflammatory diseases including inflammatory bowel disease. In this study, we assessed the therapeutic effects of MitoQ in a mouse model of experimental colitis and investigated the possible mechanisms underlying its effects on intestinal inflammation. Methods Reactive oxygen species levels and mitochondrial function were measured in blood mononuclear cells of patients with inflammatory bowel disease. The effects of MitoQ were evaluated in a dextran sulfate sodium-induced colitis mouse model. Clinical and pathological markers of disease severity and oxidative injury, and levels of inflammatory cytokines in mouse colonic tissue were measured. The effect of MitoQ on inflammatory cytokines released in the human macrophage-like cell line THP-1 was also analyzed. Results Cellular and mitochondrial reactive oxygen species levels in mononuclear cells were significantly higher in patients with inflammatory bowel disease (P <0.003, cellular reactive oxygen species; P <0.001, mitochondrial reactive oxygen species). MitoQ significantly ameliorated colitis in the dextran sulfate sodium-induced mouse model in vivo, reduced the increased oxidative stress response (malondialdehyde and 3-nitrotyrosine formation), and suppressed mitochondrial and histopathological injury by decreasing levels of inflammatory cytokines IL-1 beta and IL-18 (P <0.001 and P <0.01 respectively). By decreasing mitochondrial reactive oxygen species, MitoQ also suppressed activation of the NLRP3 inflammasome that was responsible for maturation of IL-1 beta and IL-18. In vitro studies demonstrated that MitoQ decreases IL-1 beta and IL-18 production in human THP-1 cells. Conclusion Taken together, our results suggest that MitoQ may have potential as a novel therapeutic agent for the treatment of acute phases of inflammatory bowel

  13. How mitochondrial dysfunction affects zebrafish development and cardiovascular function: an in vivo model for testing mitochondria-targeted drugs

    PubMed Central

    Pinho, Brígida R; Santos, Miguel M; Fonseca-Silva, Anabela; Valentão, Patrícia; Andrade, Paula B; Oliveira, Jorge M A

    2013-01-01

    , it evidences zebrafish's potential for in vivo efficacy or toxicity screening of ubiquinone analogues or antiparasitic mitochondria-targeted drugs. PMID:23758163

  14. How mitochondrial dysfunction affects zebrafish development and cardiovascular function: an in vivo model for testing mitochondria-targeted drugs.

    PubMed

    Pinho, Brígida R; Santos, Miguel M; Fonseca-Silva, Anabela; Valentão, Patrícia; Andrade, Paula B; Oliveira, Jorge M A

    2013-07-01

    ubiquinone analogues or antiparasitic mitochondria-targeted drugs. © 2013 The Authors. British Journal of Pharmacology © 2013 The British Pharmacological Society.

  15. Transport and metabolism of MitoQ10, a mitochondria-targeted antioxidant, in Caco-2 cell monolayers.

    PubMed

    Li, Yan; Fawcett, J Paul; Zhang, Hu; Tucker, Ian G

    2007-04-01

    Mitoquinone (MitoQ(10) mesylate) is a mitochondria-targeted antioxidant formulated for oral administration in the treatment of neurodegenerative diseases. We have investigated the absorption and metabolism of MitoQ(10) in Caco-2 cell monolayers. The intracellular accumulation of MitoQ(10) was 18-41% of the total amount of MitoQ(10) added. Some of the intracellular MitoQ(10) was reduced to mitoquinol and subsequently metabolized to glucuronide and sulfate conjugates. Transport of MitoQ(10) was polarized with the apparent permeability (P(app)) from basolateral (BL) to apical (AP) (P(appBL-->AP)) being >2.5-fold the P(app) from apical to basolateral (P(appAP-->BL)). In the presence of 4% bovine serum albumin on the basolateral side, the P(appAP-->BL) value increased 7-fold compared with control. The P(appBL-->AP) value decreased by 26, 31 and 61% in the presence of verapamil 100 microM, ciclosporin 10 and 30 microM, respectively, whereas the P(appAP-->BL) value increased 71% in the presence of ciclosporin 30 microM. Apical efflux of mitoquinol sulfate and mitoquinol glucuronide conjugates was significantly decreased by ciclosporin 30 microM and the breast cancer receptor protein (BCRP) inhibitor, reserpine 25 microM, respectively. These results suggested that the bioavailability of MitoQ(10) may be limited by intracellular metabolism and the action of P-glycoprotein and BCRP. However, the dramatic increase in absorptive P(app) in the presence of bovine serum albumin on the receiver side suggests these barrier functions may be less significant in-vivo.

  16. Inactivation of renal mitochondrial respiratory complexes and manganese superoxide dismutase during sepsis: mitochondria-targeted antioxidant mitigates injury.

    PubMed

    Patil, Naeem K; Parajuli, Nirmala; MacMillan-Crow, Lee Ann; Mayeux, Philip R

    2014-04-01

    Acute kidney injury (AKI) is a complication of sepsis and leads to a high mortality rate. Human and animal studies suggest that mitochondrial dysfunction plays an important role in sepsis-induced multi-organ failure; however, the specific mitochondrial targets damaged during sepsis remain elusive. We used a clinically relevant cecal ligation and puncture (CLP) murine model of sepsis and assessed renal mitochondrial function using high-resolution respirometry, renal microcirculation using intravital microscopy, and renal function. CLP caused a time-dependent decrease in mitochondrial complex I and II/III respiration and reduced ATP. By 4 h after CLP, activity of manganese superoxide dismutase (MnSOD) was decreased by 50% and inhibition was sustained through 36 h. These events were associated with increased mitochondrial superoxide generation. We then evaluated whether the mitochondria-targeted antioxidant Mito-TEMPO could reverse renal mitochondrial dysfunction and attenuate sepsis-induced AKI. Mito-TEMPO (10 mg/kg) given at 6 h post-CLP decreased mitochondrial superoxide levels, protected complex I and II/III respiration, and restored MnSOD activity by 18 h. Mito-TEMPO also improved renal microcirculation and glomerular filtration rate. Importantly, even delayed therapy with a single dose of Mito-TEMPO significantly increased 96-h survival rate from 40% in untreated septic mice to 80%. Thus, sepsis causes sustained inactivation of three mitochondrial targets that can lead to increased mitochondrial superoxide. Importantly, even delayed therapy with Mito-TEMPO alleviated kidney injury, suggesting that it may be a promising approach to treat septic AKI.

  17. Mitochondria-targeted ROS scavenger improves post-ischemic recovery of cardiac function and attenuates mitochondrial abnormalities in aged rats.

    PubMed

    Escobales, Nelson; Nuñez, Rebeca E; Jang, Sehwan; Parodi-Rullan, Rebecca; Ayala-Peña, Sylvette; Sacher, Joshua R; Skoda, Erin M; Wipf, Peter; Frontera, Walter; Javadov, Sabzali

    2014-12-01

    Mitochondria-generated reactive oxygen species (ROS) play a crucial role in the pathogenesis of aging and age-associated diseases. In this study, we evaluated the effects of XJB-5-131 (XJB), a mitochondria-targeted ROS and electron scavenger, on cardiac resistance to ischemia-reperfusion (IR)-induced oxidative stress in aged rats. Male adult (5-month old, n=17) and aged (29-month old, n=19) Fischer Brown Norway (F344/BN) rats were randomly assigned to the following groups: adult (A), adult+XJB (AX), aged (O), and aged+XJB (OX). XJB was administered 3 times per week (3mg/kg body weight, IP) for four weeks. At the end of the treatment period, cardiac function was continuously monitored in excised hearts using the Langendorff technique for 30 min, followed by 20 min of global ischemia, and 60-min reperfusion. XJB improved post-ischemic recovery of aged hearts, as evidenced by greater left ventricular developed-pressures and rate-pressure products than the untreated, aged-matched group. The state 3 respiration rates at complexes I, II and IV of mitochondria isolated from XJB-treated aged hearts were 57% (P<0.05), 25% (P<0.05) and 28% (P<0.05), respectively, higher than controls. Ca(2+)-induced swelling, an indicator of permeability transition pore opening, was reduced in the mitochondria of XJB-treated aged rats. In addition, XJB significantly attenuated the H2O2-induced depolarization of the mitochondrial inner membrane as well as the total and mitochondrial ROS levels in cultured cardiomyocytes. This study underlines the importance of mitochondrial ROS in aging-induced cardiac dysfunction and suggests that targeting mitochondrial ROS may be an effective therapeutic approach to protect the aged heart against IR injury.

  18. A long-linker conjugate of fluorescein and triphenylphosphonium as mitochondria-targeted uncoupler and fluorescent neuro- and nephroprotector.

    PubMed

    Antonenko, Yuri N; Denisov, Stepan S; Silachev, Denis N; Khailova, Ljudmila S; Jankauskas, Stanislovas S; Rokitskaya, Tatyana I; Danilina, Tatyana I; Kotova, Elena A; Korshunova, Galina A; Plotnikov, Egor Y; Zorov, Dmitry B

    2016-11-01

    Limited uncoupling of oxidative phosphorylation is known to be beneficial in various laboratory models of diseases. Linking a triphenyl-phosphonium cation to fluorescein through a decyl (C10) spacer yields a fluorescent uncoupler, coined mitoFluo, that selectively accumulates in energized mitochondria (Denisov et al., Chem.Commun. 2014). Proton-transport activity of mitoFluo was tested in liposomes reconstituted with bacteriorhodopsin. To examine the uncoupling action on mitochondria, we monitored mitochondrial membrane potential in parallel with oxygen consumption. Neuro- and nephroprotecting activity was detected by a limb-placing test and a kidney ischemia/reperfusion protocol, respectively. We compared mitoFluo properties with those of its newly synthesized analog having a short (butyl) spacer (C4-mitoFluo). MitoFluo, but not C4-mitoFluo, caused collapse of mitochondrial membrane potential resulting in stimulation of mitochondrial respiration. The dramatic difference in the uncoupling activity of mitoFluo and C4-mitoFluo was in line with the difference in their protonophoric activity on a lipid membrane. The accumulation of mitoFluo in mitochondria was more pronounced than that of C4-mitoFluo. MitoFluo decreased the rate of ROS production in mitochondria. MitoFluo was effective in preventing consequences of brain trauma in rats: it suppressed trauma-induced brain swelling and reduced a neurological deficit. Besides, mitoFluo attenuated acute kidney injury after ischemia/reperfusion in rats. A long alkyl linker was proved mandatory for mitoFluo to be a mitochondria- targeted uncoupler. MitoFluo showed high protective efficacy in certain models of oxidative stress-related diseases. MitoFluo is a candidate for developing therapeutic and fluorescence imaging agents to treat brain and kidney pathologies. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Assessment of H2S in vivo using the newly developed mitochondria-targeted mass spectrometry probe MitoA.

    PubMed

    Arndt, Sabine; Baeza-Garza, Carlos D; Logan, Angela; Rosa, Tiziana; Wedmann, Rudolf; Prime, Tracy A; Martin, Jack L; Saeb-Parsy, Kourosh; Krieg, Thomas; Filipovic, Milos R; Hartley, Richard C; Murphy, Michael P

    2017-05-12

    Hydrogen sulfide (H2S) is produced endogenously in vivo and has multiple effects on signaling pathways and cell function. Mitochondria can be both an H2S source and sink, and many of the biological effects of H2S relate to its interactions with mitochondria. However, the significance of mitochondrial H2S is uncertain, in part due to the difficulty of assessing changes in its concentration in vivo Although a number of fluorescent H2S probes have been developed these are best suited to cells in culture and cannot be used in vivo To address this unmet need we have developed a mitochondria-targeted H2S probe, MitoA, which can be used to assess relative changes in mitochondrial H2S levels in vivo MitoA comprises a lipophilic triphenylphosphonium (TPP) cation coupled to an aryl azide. The TPP cation leads to the accumulation of MitoA inside mitochondria within tissues in vivo There, the aryl azido group reacts with H2S to form an aryl amine (MitoN). The extent of conversion of MitoA to MitoN thus gives an indication of the levels of mitochondrial H2S in vivo Both compounds can be detected sensitively by liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis of the tissues, and quantified relative to deuterated internal standards. Here we describe the synthesis and characterization of MitoA and show that it can be used to assess changes in mitochondrial H2S levels in vivo As a proof of principle we used MitoA to show that H2S levels increase in vivo during myocardial ischemia. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Assessment of H2S in vivo using the newly developed mitochondria-targeted mass spectrometry probe MitoA

    PubMed Central

    Arndt, Sabine; Baeza-Garza, Carlos D.; Logan, Angela; Rosa, Tiziana; Wedmann, Rudolf; Prime, Tracy A.; Martin, Jack L.; Saeb-Parsy, Kourosh; Krieg, Thomas; Filipovic, Milos R.; Hartley, Richard C.; Murphy, Michael P.

    2017-01-01

    Hydrogen sulfide (H2S) is produced endogenously in vivo and has multiple effects on signaling pathways and cell function. Mitochondria can be both an H2S source and sink, and many of the biological effects of H2S relate to its interactions with mitochondria. However, the significance of mitochondrial H2S is uncertain, in part due to the difficulty of assessing changes in its concentration in vivo. Although a number of fluorescent H2S probes have been developed these are best suited to cells in culture and cannot be used in vivo. To address this unmet need we have developed a mitochondria-targeted H2S probe, MitoA, which can be used to assess relative changes in mitochondrial H2S levels in vivo. MitoA comprises a lipophilic triphenylphosphonium (TPP) cation coupled to an aryl azide. The TPP cation leads to the accumulation of MitoA inside mitochondria within tissues in vivo. There, the aryl azido group reacts with H2S to form an aryl amine (MitoN). The extent of conversion of MitoA to MitoN thus gives an indication of the levels of mitochondrial H2S in vivo. Both compounds can be detected sensitively by liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis of the tissues, and quantified relative to deuterated internal standards. Here we describe the synthesis and characterization of MitoA and show that it can be used to assess changes in mitochondrial H2S levels in vivo. As a proof of principle we used MitoA to show that H2S levels increase in vivo during myocardial ischemia. PMID:28320864

  1. Ciprofloxacin-nitroxide hybrids with potential for biofilm control.

    PubMed

    Verderosa, Anthony D; de la Fuente-Núñez, César; Mansour, Sarah C; Cao, Jicong; Lu, Timothy K; Hancock, Robert E W; Fairfull-Smith, Kathryn E

    2017-09-29

    As bacterial biofilms display extreme tolerance to conventional antibiotic treatments, it has become imperative to develop new antibacterial strategies with alternative mechanisms of action. Herein, we report the synthesis of a series of ciprofloxacin-nitroxide conjugates and their corresponding methoxyamine derivatives in high yield. This was achieved by linking various nitroxides or methoxyamines to the secondary amine of the piperazine ring of ciprofloxacin using amide bond coupling. Biological evaluation of the prepared compounds on preformed P. aeruginosa biofilms in flow cells revealed substantial dispersal with ciprofloxacin-nitroxide hybrid 25, and virtually complete killing and removal (94%) of established biofilms in the presence of ciprofloxacin-nitroxide hybrid 27. Compounds 25-28 were shown to be non-toxic in both human embryonic kidney 293 (HEK 293) cells and human muscle rhabdomyosarcoma (RD) cells at concentrations up to 40 μM. Significantly, these hybrids demonstrate the potential of antimicrobial-nitroxide agents to overcome the resistance of biofilms to antimicrobials via stimulation of biofilm dispersal or through direct cell killing. Crown Copyright © 2017. Published by Elsevier Masson SAS. All rights reserved.

  2. Modulating mtDNA heteroplasmy by mitochondria-targeted restriction endonucleases in a ‘differential multiple cleavage-site’ model

    PubMed Central

    Bacman, SR; Williams, SL; Hernandez, D; Moraes, CT

    2009-01-01

    The ability to manipulate mitochondrial DNA (mtDNA) heteroplasmy would provide a powerful tool to treat mitochondrial diseases. Recent studies showed that mitochondria-targeted restriction endonucleases can modify mtDNA heteroplasmy in a predictable and efficient manner if it recognizes a single site in the mutant mtDNA. However, the applicability of such model is limited to mutations that create a novel cleavage site, not present in the wild-type mtDNA. We attempted to extend this approach to a ‘differential multiple cleavage site’ model, where an mtDNA mutation creates an extra restriction site to the ones normally present in the wild-type mtDNA. Taking advantage of a heteroplasmic mouse model harboring two haplotypes of mtDNA (NZB/BALB) and using adenovirus as a gene vector, we delivered a mitochondria-targeted Scal restriction endonuclease to different mouse tissues. Scal recognizes five sites in the NZB mtDNA but only three in BALB mtDNA. Our results showed that changes in mtDNA heteroplasmy were obtained by the expression of mitochondria-targeted ScaI in both liver, after intravenous injection, and in skeletal muscle, after intramuscular injection. Although mtDNA depletion was an undesirable side effect, our data suggest that under a regulated expression system, mtDNA depletion could be minimized and restriction endonucleases recognizing multiple sites could have a potential for therapeutic use. PMID:17597792

  3. The Development of Nitroxide Based Coatings for Biofilm Remediation- 154020

    DTIC Science & Technology

    2017-06-05

    AFRL-AFOSR-JP-TR-2017-0048 The Development of Nitroxide Based Coatings for Biofilm Remediation Kathryn Fairfull-Smith QUEENSLAND UNIVERSITY OF...does not display a currently valid OMB control number. PLEASE DO NOT RETURN YOUR FORM TO THE ABOVE ORGANIZATION . 1. REPORT DATE (DD-MM-YYYY)      19-06...2017 2. REPORT TYPE Final 3. DATES COVERED (From - To) 08 Sep 2015 to 07 Mar 2017 4. TITLE AND SUBTITLE The Development of Nitroxide Based Coatings

  4. The β-phosphorus hyperfine coupling constant in nitroxides: 6. Solvent effects in non-cyclic nitroxides.

    PubMed

    Audran, Gérard; Bosco, Lionel; Nkolo, Paulin; Bikanga, Raphael; Brémond, Paul; Butscher, Teddy; Marque, Sylvain R A

    2016-04-12

    In two recent articles (Org. Biomol. Chem., 2015 and 2016), we showed that changes in the phosphorus hyperfine coupling constant aP at position β in β-phosphorylated nitroxides can be dramatic. Such changes were applied to the titration of water in organic solvents and conversely of organic solvents in water. One of the molecules tested was a non-cyclic nitroxide meaning that a thorough investigation of the solvent effect on the EPR hyperfine coupling constant is timely due. In this article, we show that the aP of persistent non-cyclic β-phosphorylated nitroxides decrease with the normalized polarity Reichardt's constant E(N)T. The Koppel-Palm and Kalmet-Abboud-Taft relationships were applied to gain deeper insight into the effects influencing aN and aP: polarity/polarizability, hydrogen bond donor properties, and the structuredness of the cybotactic region.

  5. Mitochondria-targeted Ogg1 and aconitase-2 prevent oxidant-induced mitochondrial DNA damage in alveolar epithelial cells.

    PubMed

    Kim, Seok-Jo; Cheresh, Paul; Williams, David; Cheng, Yuan; Ridge, Karen; Schumacker, Paul T; Weitzman, Sigmund; Bohr, Vilhelm A; Kamp, David W

    2014-02-28

    Mitochondria-targeted human 8-oxoguanine DNA glycosylase (mt-hOgg1) and aconitase-2 (Aco-2) each reduce oxidant-induced alveolar epithelial cell (AEC) apoptosis, but it is unclear whether protection occurs by preventing AEC mitochondrial DNA (mtDNA) damage. Using quantitative PCR-based measurements of mitochondrial and nuclear DNA damage, mtDNA damage was preferentially noted in AEC after exposure to oxidative stress (e.g. amosite asbestos (5-25 μg/cm(2)) or H2O2 (100-250 μM)) for 24 h. Overexpression of wild-type mt-hOgg1 or mt-long α/β 317-323 hOgg1 mutant incapable of DNA repair (mt-hOgg1-Mut) each blocked A549 cell oxidant-induced mtDNA damage, mitochondrial p53 translocation, and intrinsic apoptosis as assessed by DNA fragmentation and cleaved caspase-9. In contrast, compared with controls, knockdown of Ogg1 (using Ogg1 shRNA in A549 cells or primary alveolar type 2 cells from ogg1(-/-) mice) augmented mtDNA lesions and intrinsic apoptosis at base line, and these effects were increased further after exposure to oxidative stress. Notably, overexpression of Aco-2 reduced oxidant-induced mtDNA lesions, mitochondrial p53 translocation, and apoptosis, whereas siRNA for Aco-2 (siAco-2) enhanced mtDNA damage, mitochondrial p53 translocation, and apoptosis. Finally, siAco-2 attenuated the protective effects of mt-hOgg1-Mut but not wild-type mt-hOgg1 against oxidant-induced mtDNA damage and apoptosis. Collectively, these data demonstrate a novel role for mt-hOgg1 and Aco-2 in preserving AEC mtDNA integrity, thereby preventing oxidant-induced mitochondrial dysfunction, p53 mitochondrial translocation, and intrinsic apoptosis. Furthermore, mt-hOgg1 chaperoning of Aco-2 in preventing oxidant-mediated mtDNA damage and apoptosis may afford an innovative target for the molecular events underlying oxidant-induced toxicity.

  6. Nitroxide stable radicals protect beating cardiomyocytes against oxidative damage

    SciTech Connect

    Samuni, A.; Winkelsberg, D.; Pinson, A.; Hahn, S.M.; Mitchell, J.B.; Russo, A. )

    1991-05-01

    The protective effect of stable nitroxide radicals against oxidative damage was studied using cardiomyocyte cultures obtained from newborn rats. Monolayered cardiomyocytes were exposed to H{sub 2}O{sub 2} and the effect on spontaneous beating and leakage of LDH was determined. Hydrogen peroxide irreversibly blocked rhythmic beating and resulted in a significant membrane injury as shown by release of LDH. The injury was prevented by catalase which removes H{sub 2}O{sub 2} and by cell-permeable, metal-chelating agents such as desferrioxamine or bipyridine. In contrast, reagents which are excluded from the cell such as superoxide dismutase or DTPA did not protect the cells against H{sub 2}O{sub 2}. Five- and six-membered ring, stable nitroxide radicals which have previously been shown to chemically act as low-molecular weight, membrane-permeable, SOD-mimetic compounds provided full protection. The nitroxides prevented leakage of LDH and preserved normal cardiomyocyte contractility, presumably by intercepting intracellular O{sub 2}-radicals. Alternatively, protection may result through nitroxides reacting with reduced transition metal ions or by detoxifying secondary organic radicals.

  7. Preparation of Some Homologous TEMPO Nitroxides and Oxoammonium Salts; Notes on the NMR Spectroscopy of Nitroxide Free Radicals; Observed Radical Nature of Oxoammonium Salt Solutions Containing Trace Amounts of Corresponding Nitroxides in an Equilibrium Relationship.

    PubMed

    Bobbitt, James M; Eddy, Nicholas A; Cady, Clyde X; Jin, Jing; Gascon, Jose A; Gelpí-Dominguez, Svetlana; Zakrzewski, Jerzy; Morton, Martha D

    2017-09-06

    Three new homologous TEMPO oxoammonium salts and three homologous nitroxide radicals have been prepared and characterized. The oxidation properties of the salts have been explored. The direct (13)C NMR and EPR spectra of the nitroxide free radicals and the oxoammonium salts, along with TEMPO and its oxoammonium salt, have been successfully measured with little peak broadening of the NMR signals. In the spectra of all ten compounds (nitroxides and corresponding oxoammonium salts), the carbons in the 2,2,6,6-tetramethylpiperidine core do not appear, implying paramagnetic properties. This unpredicted overall paramagnetism in the oxoammonium salt solutions is explained by a redox equilibrium as shown between oxoammonium salts and trace amounts of corresponding nitroxide. This equilibrium is confirmed by electron interchange reactions between nitroxides with an N-acetyl substituent and oxoammonium salts with longer acyl side chains.

  8. Nitroxides as redox probes of melanins: dark-induced and photoinduced changes in redox equilibria

    SciTech Connect

    Sarna, T.; Korytowski, W.; Sealy, R.C.

    1985-05-15

    The interaction of nitroxide free radicals and their reduced products (hydroxylamines) with synthetic and natural melanins has been studied. Electron spin resonance spectroscopy was used to measure changes in radical concentration in the dark and during irradiation with visible or uv light. Some reduction of nitroxide occurs in the dark, and is reversible: the nitroxide can be completely regenerated by the one-electron oxidant ferricyanide. The kinetics of the process depend strongly on radical charge and pH. For positively charged nitroxides the rate is much faster than for either neutral or anionic radicals. At pH 10 the rate is about 20 times faster than at pH 5. Oxidation of hydroxylamine also can occur so that a redox equilibrium is established. The equilibrium constant has been estimated for the reaction between a nitroxide and melanin from autoxidation of 3,4-dihydroxyphenylalanine. Results are also dependent upon the type of melanin used and chemical modification (oxidation or reduction) of the melanin. Redox equilibria are altered during irradiation with either visible or uv light. Rapid oxidation of hydroxylamine to nitroxide is apparent, together with a slower reduction of nitroxide. Action spectra for these processes are related to those for melanin radical production and oxygen consumption in nitroxide-free melanin systems. Reduction of nitroxide is inhibited by oxygen, suggesting a competition between nitroxide and oxygen for photoinduced reducing equivalents.

  9. Cardioprotective Effects of Mitochondria-Targeted Peptide SBT-20 in two Different Models of Rat Ischemia/Reperfusion.

    PubMed

    Dai, Wangde; Cheung, Elissa; Alleman, Rick J; Perry, Justin B; Allen, Mitchell E; Brown, David A; Kloner, Robert A

    2016-12-01

    Dysfunctional mitochondria are considered to be the major source of intracellular reactive oxygen species and play a central role in the pathophysiology of myocardial ischemia/reperfusion. This study sought to determine effects of mitochondria-targeted cytoprotective peptide SBT-20 on myocardial infarct size in two different models of ischemia/reperfusion. For in vivo studies, anesthetized Sprague Dawley rats were subjected to 30 min of coronary artery occlusion followed by 3 h of reperfusion. Rats received saline (control), low dose SBT-20 (0.3 mg/kg/h) or high dose SBT-20 (3 mg/kg/h) treatment (n = 15 rats in each group). Saline or SBT-20 were delivered into the jugular vein starting 5 min after coronary artery occlusion and were continued for one hour post coronary artery reperfusion. Body temperature, heart rate and blood pressure were monitored during the procedure. At the end of 3 h reperfusion, the ischemic risk area, no-reflow area, and infarct size were measured. In separate in vitro studies, isolated rat hearts were exposed to 20 min global ischemia, followed by SBT-20 administration (1 μM) or no SBT-20 (control) throughout the 2 h reperfusion. In vitro studies were conducted in cells and heart mitochondria to ascertain the mitochondrial effects of SBT-20 on mitochondrial respiration and reactive oxygen species production. In the in vivo study, the ischemic risk areas (as a percentage of the left ventricle) were similar among the saline (49.5 ± 2.3 %), low dose SBT-20 (48.6 ± 2.1 %), and high dose SBT-20 groups (48.7 ± 3.0 %). Treatment with SBT-20 significantly reduced infarct size ( as a percentage of risk area) in low dose (62.1 ± 4.4 %) and high dose (64.0 ± 4.9 %) compared with saline treatment (77.6 ± 2.6 %, p = 0.001 for both doses). There was no difference in infarct size between low and high dose SBT-20 treatment. The no-reflow areas (as a percentage of the risk area) were comparable among the saline (23.9

  10. Mitochondria-targeted heme oxygenase-1 induces oxidative stress and mitochondrial dysfunction in macrophages, kidney fibroblasts and in chronic alcohol hepatotoxicity.

    PubMed

    Bansal, Seema; Biswas, Gopa; Avadhani, Narayan G

    2014-01-01

    The inducible form of Heme Oxygenase-1 (HO-1), a major endoplasmic reticulum (ER) associated heme protein, is known to play important roles in protection against oxidative and chemical stress by degrading free heme released from degradation of heme proteins. In this study we show that induced expression of HO-1 by subjecting macrophage RAW-264.7 cells to chemical or physiological hypoxia resulted in significant translocation of HO-1 protein to mitochondria. Transient transfection of COS-7 cells with cloned cDNA also resulted in mitochondrial translocation of HO-1. Deletion of N-terminal ER targeting domain increased mitochondrial translocation under the transient transfection conditions. Mitochondrial localization of both intact HO-1 and N-terminal truncated HO-1 caused loss of heme aa-3 and cytochrome c oxidase (CcO) activity in COS-7 cells. The truncated protein, which localizes to mitochondria at higher levels, induced substantially steeper loss of CcO activity and reduced heme aa3 content. Furthermore, cells expressing mitochondria targeted HO-1 also induced higher ROS production. Consistent with dysfunctional state of mitochondria induced by HO-1, the mitochondrial recruitment of autophagy markers LC-3 and Drp-1 was also increased in these cells. Chronic ethanol feeding in rats also caused an increase in mitochondrial HO-1 and decrease in CcO activity. These results show that as opposed to the protective effect of the ER associated HO-1, mitochondria targeted HO-1 under normoxic conditions induces mitochondrial dysfunction.

  11. Alzheimer's disease-like pathology in senescence-accelerated OXYS rats can be partially retarded with mitochondria-targeted antioxidant SkQ1.

    PubMed

    Stefanova, Natalia A; Muraleva, Natalia A; Skulachev, Vladimir P; Kolosova, Nataliya G

    2014-01-01

    We previously showed that mitochondria-targeted antioxidant SkQ1 (plastoquinonyl-decyltriphenylphosphonium) at nanomolar concentrations is capable of preventing and slowing down some cerebral dysfunctions in accelerated-senescence OXYS rats. Here we demonstrate that OXYS rats develop behavior, learning, and memory deficits against a background of neurodegeneration signs detected by magnetic resonance tomography and amyloid-β (Aβ) pathology similar to those seen in Alzheimer's disease (AD). Long-term treatment with SkQ1 (250 nmol/kg body weight daily from the age of 1.5 to 23 months) reduced the age-related alterations in behavior and spatial memory deficit in Morris water maze in OXYS and Wistar rats. Furthermore, this is the first report that SkQ1 treatment slows down pathological accumulation of AβPP, Aβ, and hyperphosphorylation of tau-protein in OXYS rats, as well as age-dependent changes in healthy Wistar rats. Our results support the possibility of using the OXYS strain as a rat model of AD-like pathology. It seems probable that the mitochondria-targeted antioxidant SkQ1 can be a good prophylactic strategy to maintain brain health and to treat AD.

  12. The Mitochondria-Targeted Antioxidant MitoQ Prevents Loss of Spatial Memory Retention and Early Neuropathology in a Transgenic Mouse Model of Alzheimer’s Disease

    PubMed Central

    McManus, Meagan J.; Murphy, Michael P.; Franklin, James L.

    2012-01-01

    Considerable evidence suggests that mitochondrial dysfunction and oxidative stress contribute to the progression of Alzheimer’s disease (AD). We examined the ability of the novel mitochondria-targeted antioxidant MitoQ (mitoquinone mesylate: [10-(4,5-dimethoxy-2-methyl-3,6-dioxo-1,4-cycloheexadienlyl) decyl triphenylphosphonium methanesulfonate]) to prevent AD-like pathology in mouse cortical neurons in cell culture and in a triple transgenic mouse model of AD (3xTg-AD). MitoQ attenuated β-amyloid (Aβ)-induced neurotoxicity in cortical neurons and also prevented increased production of reactive species and loss of mitochondrial membrane potential (Δψm) in them. To determine whether the mitochondrial protection conferred by MitoQ was sufficient to prevent the emergence of AD-like neuropathology in vivo, we treated young female 3xTg-AD mice with MitoQ for 5 months and analyzed the effect on the progression of AD-like pathologies. Our results show that MitoQ prevented cognitive decline in these mice as well as oxidative stress, Aβ accumulation, astrogliosis, synaptic loss, and caspase activation in their brains. The work presented herein suggests a central role for mitochondria in neurodegeneration and provides evidence supporting the use of mitochondria-targeted therapeutics in diseases involving oxidative stress and metabolic failure, namely AD. PMID:22049413

  13. Mitochondria-targeted heme oxygenase-1 induces oxidative stress and mitochondrial dysfunction in macrophages, kidney fibroblasts and in chronic alcohol hepatotoxicity☆

    PubMed Central

    Bansal, Seema; Biswas, Gopa; Avadhani, Narayan G.

    2013-01-01

    The inducible form of Heme Oxygenase-1 (HO-1), a major endoplasmic reticulum (ER) associated heme protein, is known to play important roles in protection against oxidative and chemical stress by degrading free heme released from degradation of heme proteins. In this study we show that induced expression of HO-1 by subjecting macrophage RAW-264.7 cells to chemical or physiological hypoxia resulted in significant translocation of HO-1 protein to mitochondria. Transient transfection of COS-7 cells with cloned cDNA also resulted in mitochondrial translocation of HO-1. Deletion of N-terminal ER targeting domain increased mitochondrial translocation under the transient transfection conditions. Mitochondrial localization of both intact HO-1 and N-terminal truncated HO-1 caused loss of heme aa-3 and cytochrome c oxidase (CcO) activity in COS-7 cells. The truncated protein, which localizes to mitochondria at higher levels, induced substantially steeper loss of CcO activity and reduced heme aa3 content. Furthermore, cells expressing mitochondria targeted HO-1 also induced higher ROS production. Consistent with dysfunctional state of mitochondria induced by HO-1, the mitochondrial recruitment of autophagy markers LC-3 and Drp-1 was also increased in these cells. Chronic ethanol feeding in rats also caused an increase in mitochondrial HO-1 and decrease in CcO activity. These results show that as opposed to the protective effect of the ER associated HO-1, mitochondria targeted HO-1 under normoxic conditions induces mitochondrial dysfunction. PMID:24494190

  14. Synthesis and functional survey of new Tacrine analogs modified with nitroxides or their precursors

    PubMed Central

    Kálai, Tamás; Altman, Robin; Maezawa, Izumi; Balog, Mária; Morisseau, Christophe; Petrlova, Jitka; Hammock, Bruce D.; Jin, Lee-Way; Trudell, James; Voss, John C.; Hideg, Kálmán

    2014-01-01

    A series of new Tacrine analogs modified with nitroxides or pre-nitroxides on 9-amino group via methylene or piperazine spacers were synthesized; the nitroxide or its precursors were incorporated into the Tacrine scaffold. The new compounds were tested for their hydroxyl radical and peroxyl radical scavenging ability, acetyl cholinesterase inhibitor activity and protection against Aβ-induced cytotoxicity. Based on these assays, we conclude that Tacrine analogs connected to five and six-membered nitroxides via piperazine spacers (9b, 9b/HCl and 12) exhibited the best activity, providing direction for further development of additional candidates with dual functionality (anti Alzheimer’s and antioxidant). PMID:24657571

  15. Photoprotective and radioprotective properties of nitroxides and their application in magnetic resonance imaging.

    PubMed

    Lewandowski, Marcin; Gwoździński, Krzysztof

    2016-10-14

    Nitroxides are a group of stable organic radicals of low molecular weight having a nitroxyl group > N-.O, which has an unpaired electron. The presence of this group allows a nitroxide to participate in redox reactions. They serve as mimics of superoxide dismutase (SOD) and have stimulative properties towards haemoproteins with catalase-like activity. Nitroxides oxidize Fe (II) to Fe (III) preventing the Fenton and Haber-Weiss reactions. As the radicals have the ability to scavenge other free radicals. Nitroxides are not immunogenic, and mutagenic and do not show toxicity to the human cells. The review discusses the use of nitroxide in protecting cells and tissues from the effects of UVA radiation. Preliminary studies indicate that they are more effective than conventionally used vitamins C and E and UV filters. They also protect the biological material from the effects of ionizing radiation. Nitroxides protect healthy cells and simultaneously they do not protect cancer cells from ionizing radiation. The differences in the nitroxide activity are associated with conditions prevailing in the oxidizing environment of the tumor as opposed to reducing conditions in normal cells. Nitroxides can be used as contrast agents in the magnetic resonance imaging (MR). They have ability of detection of subtle changes in redox equilibrium in the tumor tissue. Application of nitroxides in MR method allow to distinguish normal and pathological state of tissue. Successful investigations using this technique were conducted in mice with colon and brain cancer.

  16. Spin-polarized nitroxide radicals in organic glasses.

    SciTech Connect

    Tarasov, V. F.; Shkrob, I. A.; Trifunac, A. D.; Chemistry

    2002-01-01

    Nonequilibrium spin polarization formed in a stable nitroxide radical, 2,2,6,6-tetramethyl-1-piperidinyloxy (Tempo) due to the occurrence of Chemically Induced Dynamic Electron Polarization (CIDEP) in photoexcited molecular complexes of this radical with 1,4-benzoquinone, 1,4-naphthaquinone, 9,10-anthraquinone, and their derivatives is observed. These complexes occur spontaneously in low-temperature organic glasses (20-70 K) upon freezing the concentrated liquid solutions. The emissive net polarization in the nitroxide radical is observed 0.1-10 {mu}s after the photoexcitation of the p-quinone moiety. No degradation of the polarized magnetic resonance signal from Tempo after >104 excitation cycles was observed. This spin polarization is shown to be mainly due to a polarization transfer from the lowest triplet state of the p-quinone. This transfer is driven by the electron spin exchange interaction between the nitroxide radical and the triplet p-quinone; it occurs simultaneously with a spin-selective electronic relaxation of the photoexcited complex. The resulting mechanism combines the features of the electron spin polarization transfer (ESPT) and radical-triplet pair mechanisms (RTPM) in liquid. A theoretical model of such a mechanism is suggested.

  17. Nitroxide pharmaceutical development for age-related degeneration and disease

    PubMed Central

    Zarling, Jacob A.; Brunt, Vienna E.; Vallerga, Anne K.; Li, Weixing; Tao, Albert; Zarling, David A.; Minson, Christopher T.

    2015-01-01

    Nitroxide small molecule agents are in development as preventative or therapeutic pharmaceutical drugs for age-related macular degeneration (AMD) and cardiovascular disease, which are two major diseases of aging. These aging diseases are associated with patient genetics, smoking, diet, oxidative stress, and chronic inflammation. Nitroxide drugs preventing aging-, smoking-, high sugar or high fat diet-, or radiation- and other environmental-induced pathophysiological conditions in aging disease are reviewed. Tempol (TP), Tempol Hydroxylamine (TP-H), and TP-H prodrug (OT-551) are evaluated in (1) non-smokers versus smokers with cutaneous microvascular dysfunction, rapidly reversed by cutaneous TP; (2) elderly cancer patients at risk for radiation-induced skin burns or hair loss, prevented by topical TP; and (3) elderly smoker or non-smoker AMD patients at risk for vision loss, prevented by daily eye drops of OT-551. The human data indicates safety and efficacy for these nitroxide drugs. Both TP and TP-H topically penetrate and function in skin or mucosa, protecting and treating radiation burns and hair loss or smoking-induced cutaneous vascular dysfunction. TP and TP-H do not penetrate the cornea, while OT-551 does effectively penetrate and travels to the back of the eye, preserving visual acuity and preserving normal and low light luminance in dry AMD smokers and non-smoker patients. Topical, oral, or injectable drug formulations are discussed. PMID:26594225

  18. Nitroxide pharmaceutical development for age-related degeneration and disease.

    PubMed

    Zarling, Jacob A; Brunt, Vienna E; Vallerga, Anne K; Li, Weixing; Tao, Albert; Zarling, David A; Minson, Christopher T

    2015-01-01

    Nitroxide small molecule agents are in development as preventative or therapeutic pharmaceutical drugs for age-related macular degeneration (AMD) and cardiovascular disease, which are two major diseases of aging. These aging diseases are associated with patient genetics, smoking, diet, oxidative stress, and chronic inflammation. Nitroxide drugs preventing aging-, smoking-, high sugar or high fat diet-, or radiation- and other environmental-induced pathophysiological conditions in aging disease are reviewed. Tempol (TP), Tempol Hydroxylamine (TP-H), and TP-H prodrug (OT-551) are evaluated in (1) non-smokers versus smokers with cutaneous microvascular dysfunction, rapidly reversed by cutaneous TP; (2) elderly cancer patients at risk for radiation-induced skin burns or hair loss, prevented by topical TP; and (3) elderly smoker or non-smoker AMD patients at risk for vision loss, prevented by daily eye drops of OT-551. The human data indicates safety and efficacy for these nitroxide drugs. Both TP and TP-H topically penetrate and function in skin or mucosa, protecting and treating radiation burns and hair loss or smoking-induced cutaneous vascular dysfunction. TP and TP-H do not penetrate the cornea, while OT-551 does effectively penetrate and travels to the back of the eye, preserving visual acuity and preserving normal and low light luminance in dry AMD smokers and non-smoker patients. Topical, oral, or injectable drug formulations are discussed.

  19. Nitroxide-Functionalized Graphene Oxide from Graphite Oxide

    PubMed Central

    Avila-Vega, Yazmin I.; Leyva-Porras, Cesar C.; Mireles, Marcela; Quevedo-López, Manuel; Macossay, Javier; Bonilla-Cruz, José

    2013-01-01

    A facile method for preparing functionalized graphene oxide single layers with nitroxide groups is reported herein. Highly oxidized graphite oxide (GO=90.6%) was obtained, slightly modifying an improved Hummer’s method. Oxoammonium salts (OS) were investigated to introduce nitroxide groups to GO, resulting in a one-step functionalization and exfoliation. The mechanisms of functionalization/exfoliation are proposed, where the oxidation of aromatic alcohols to ketone groups, and the formation of alkoxyamine species are suggested. Two kinds of functionalized graphene oxide layers (GOFT1 and GOFT2) were obtained by controlling the amount of OS added. GOFT1 and GOFT2 exhibited a high interlayer spacing (d0001 = 1.12nm), which was determined by X-ray diffraction. The presence of new chemical bonds C-N (~9.5 %) and O-O (~4.3 %) from nitroxide attached onto graphene layers were observed by X-ray photoelectron spectroscopy. Single-layers of GOFT1 were observed by HRTEM, exhibiting amorphous and crystalline zones at a 50:50 ratio; in contrast, layers of GOFT2 exhibited a fully amorphous surface. Fingerprint of GOFT1 single layers was obtained by electron diffraction at several tilts. Finally, the potential use of these materials within Nylon 6 matrices was investigated, where an unusual simultaneous increase in tensile stress, tensile strain and Young’s modulus was observed. PMID:24347671

  20. Mitochondria-targeted antioxidant promotes recovery of skeletal muscle mitochondrial function after burn trauma assessed by in vivo 31P nuclear magnetic resonance and electron paramagnetic resonance spectroscopy

    PubMed Central

    Righi, Valeria; Constantinou, Caterina; Mintzopoulos, Dionyssios; Khan, Nadeem; Mupparaju, S. P.; Rahme, Laurence G.; Swartz, Harold M.; Szeto, Hazel H.; Tompkins, Ronald G.; Tzika, A. Aria

    2013-01-01

    Burn injury causes a major systemic catabolic response that is associated with mitochondrial dysfunction in skeletal muscle. We investigated the effects of the mitochondria-targeted peptide antioxidant Szeto-Schiller 31 (SS-31) on skeletal muscle in a mouse burn model using in vivo phosphorus-31 nuclear magnetic resonance (31P NMR) spectroscopy to noninvasively measure high-energy phosphate levels; mitochondrial aconitase activity measurements that directly correlate with TCA cycle flux, as measured by gas chromatography mass spectrometry (GC-MS); and electron paramagnetic resonance (EPR) to assess oxidative stress. At 6 h postburn, the oxidative ATP synthesis rate was increased 5-fold in burned mice given a single dose of SS-31 relative to untreated burned mice (P=0.002). Furthermore, SS-31 administration in burned animals decreased mitochondrial aconitase activity back to control levels. EPR revealed a recovery in redox status of the SS-31-treated burn group compared to the untreated burn group (P<0.05). Our multidisciplinary convergent results suggest that SS-31 promotes recovery of mitochondrial function after burn injury by increasing ATP synthesis rate, improving mitochondrial redox status, and restoring mitochondrial coupling. These findings suggest use of noninvasive in vivo NMR and complementary EPR offers an approach to monitor the effectiveness of mitochondrial protective agents in alleviating burn injury symptoms.—Righi, V., Constantinou, C., Mintzopoulos, D., Khan, N., Mupparaju, S. P., Rahme, L. G., Swartz, H. M., Szeto, H. H., Tompkins, R. G., and Tzika, A. A. Mitochondria-targeted antioxidant promotes recovery of skeletal muscle mitochondrial function after burn trauma assessed by in vivo 31P nuclear magnetic resonance and electron paramagnetic resonance spectroscopy. PMID:23482635

  1. The mitochondria-targeted anti-oxidant MitoQ reduces aspects of mitochondrial fission in the 6-OHDA cell model of Parkinson's disease.

    PubMed

    Solesio, María E; Prime, Tracy A; Logan, Angela; Murphy, Michael P; Del Mar Arroyo-Jimenez, María; Jordán, Joaquín; Galindo, María F

    2013-01-01

    Parkinson's disease (PD) is a neurodegenerative disorder for which available treatments provide symptom relief but do not stop disease progression. Mitochondria, and in particular mitochondrial dynamics, have been postulated as plausible pharmacological targets. Mitochondria-targeted antioxidants have been developed to prevent mitochondrial oxidative damage, and to alter the involvement of reactive oxygen species (ROS) in signaling pathways. In this study, we have dissected the effect of MitoQ, which is produced by covalent attachment of ubiquinone to a triphenylphosphonium lipophilic cation by a ten carbon alkyl chain. MitoQ was tested in an in vitro PD model which involves addition of 6-hydroxydopamine (6-OHDA) to SH-SY5Y cell cultures. At sublethal concentrations of 50μM, 6-OHDA did not induce increases in protein carbonyl, mitochondrial lipid peroxidation or mitochondrial DNA damage. However, after 3h of treatment, 6-OHDA disrupts the mitochondrial morphology and activates the machinery of mitochondrial fission, but not fusion. Addition of 6-OHDA did not increase the levels of fission 1, mitofusins 1 and 2 or optic atrophy 1 proteins, but does lead to the translocation of dynamin related protein 1 from the cytosol to the mitochondria. Pre-treatment with MitoQ (50nM, 30min) results in the inhibition of the mitochondrial translocation of Drp1. Furthermore, MitoQ also inhibited the translocation of the pro-apoptotic protein Bax to the mitochondria. These findings provide mechanistic evidence for a role for redox events contributing to mitochondrial fission and suggest the potential of mitochondria-targeted therapeutics in diseases that involve mitochondrial fragmentation due to oxidative stress.

  2. Mitochondria-targeted esculetin alleviates mitochondrial dysfunction by AMPK-mediated nitric oxide and SIRT3 regulation in endothelial cells: potential implications in atherosclerosis.

    PubMed

    Karnewar, Santosh; Vasamsetti, Sathish Babu; Gopoju, Raja; Kanugula, Anantha Koteswararao; Ganji, Sai Krishna; Prabhakar, Sripadi; Rangaraj, Nandini; Tupperwar, Nitin; Kumar, Jerald Mahesh; Kotamraju, Srigiridhar

    2016-04-11

    Mitochondria-targeted compounds are emerging as a new class of drugs that can potentially alter the pathophysiology of those diseases where mitochondrial dysfunction plays a critical role. We have synthesized a novel mitochondria-targeted esculetin (Mito-Esc) with an aim to investigate its effect during oxidative stress-induced endothelial cell death and angiotensin (Ang)-II-induced atherosclerosis in ApoE(-/-) mice. Mito-Esc but not natural esculetin treatment significantly inhibited H2O2- and Ang-II-induced cell death in human aortic endothelial cells by enhancing NO production via AMPK-mediated eNOS phosphorylation. While L-NAME (NOS inhibitor) significantly abrogated Mito-Esc-mediated protective effects, Compound c (inhibitor of AMPK) significantly decreased Mito-Esc-mediated increase in NO production. Notably, Mito-Esc promoted mitochondrial biogenesis by enhancing SIRT3 expression through AMPK activation; and restored H2O2-induced inhibition of mitochondrial respiration. siSIRT3 treatment not only completely reversed Mito-Esc-mediated mitochondrial biogenetic marker expressions but also caused endothelial cell death. Furthermore, Mito-Esc administration to ApoE(-/-) mice greatly alleviated Ang-II-induced atheromatous plaque formation, monocyte infiltration and serum pro-inflammatory cytokines levels. We conclude that Mito-Esc is preferentially taken up by the mitochondria and preserves endothelial cell survival during oxidative stress by modulating NO generation via AMPK. Also, Mito-Esc-induced SIRT3 plays a pivotal role in mediating mitochondrial biogenesis and perhaps contributes to its anti-atherogenic effects.

  3. Mitochondria-targeted esculetin alleviates mitochondrial dysfunction by AMPK-mediated nitric oxide and SIRT3 regulation in endothelial cells: potential implications in atherosclerosis

    PubMed Central

    Karnewar, Santosh; Vasamsetti, Sathish Babu; Gopoju, Raja; Kanugula, Anantha Koteswararao; Ganji, Sai Krishna; Prabhakar, Sripadi; Rangaraj, Nandini; Tupperwar, Nitin; Kumar, Jerald Mahesh; Kotamraju, Srigiridhar

    2016-01-01

    Mitochondria-targeted compounds are emerging as a new class of drugs that can potentially alter the pathophysiology of those diseases where mitochondrial dysfunction plays a critical role. We have synthesized a novel mitochondria-targeted esculetin (Mito-Esc) with an aim to investigate its effect during oxidative stress-induced endothelial cell death and angiotensin (Ang)-II-induced atherosclerosis in ApoE−/− mice. Mito-Esc but not natural esculetin treatment significantly inhibited H2O2- and Ang-II-induced cell death in human aortic endothelial cells by enhancing NO production via AMPK-mediated eNOS phosphorylation. While L-NAME (NOS inhibitor) significantly abrogated Mito-Esc-mediated protective effects, Compound c (inhibitor of AMPK) significantly decreased Mito-Esc-mediated increase in NO production. Notably, Mito-Esc promoted mitochondrial biogenesis by enhancing SIRT3 expression through AMPK activation; and restored H2O2-induced inhibition of mitochondrial respiration. siSIRT3 treatment not only completely reversed Mito-Esc-mediated mitochondrial biogenetic marker expressions but also caused endothelial cell death. Furthermore, Mito-Esc administration to ApoE−/− mice greatly alleviated Ang-II-induced atheromatous plaque formation, monocyte infiltration and serum pro-inflammatory cytokines levels. We conclude that Mito-Esc is preferentially taken up by the mitochondria and preserves endothelial cell survival during oxidative stress by modulating NO generation via AMPK. Also, Mito-Esc-induced SIRT3 plays a pivotal role in mediating mitochondrial biogenesis and perhaps contributes to its anti-atherogenic effects. PMID:27063143

  4. [Inhibitory effect of human mitochondria-targeted MPG recombinant on proliferation of human non-small cell lung cancer multidrug-resistant cell line A549/DDP].

    PubMed

    Yu, Shi-Cang; Qian, Gui-Sheng; Li, Yu-Ying; Lu, Wei-Zhong; Li, Jin; Huang, Gui-Jun

    2006-04-01

    Multidrug resistance is the key obstacle to the improvement of chemotherapy effect of lung cancer. This study was to construct eukaryotic expression vector of human mitochondria-targeted N-methylpurine DNA glycosylase (MPG), and explore its inhibitory effect on proliferation of human non-small cell lung cancer multidrug-resistant cell line A549/DDP. Manganese-superoxide dismutase mitochondria-targeted sequence-MPG fusion gene (mito-MPG) was constructed through splicing by overlap extension (SOE). Recombinant eukaryotic expression vector pCMV-Script/mito-MPG was constructed by molecule-cloning technique, and then transfected into A549/DDP cells. In the stably transfected cells which were screened out by G418, the expression of mito-MPG mRNA was detected by reverse transcription-polymerase chain reaction (RT-PCR); its expression in separated and purified mitochondria was detected by Western blot. The proliferation of A549/DDP cells was detected by trypan blue exclusion trial. Cell cycle distribution was analyzed by flow cytometry. The mito-MPG fusion gene was confirmed by DNA sequencing,the recombinant pCMV-Script/mito-MPG was confirmed by restrictive endonuclease digestion and DNA sequencing. mito-MPG mRNA and protein were detected in the cells transfected with pCMV-Script/mito-MPG (MPG group), but not in the cells transfected with pCMV-Script (P group) and untransfected cells (C group). The cell double time were 72.6 h in C group, 73.5 h in P group, and 98.9 h in MPG group. Cell cycle blockage and subdiploid peak were found in MPG group. The proliferation indexes were 51.3% in C group, 54.3% in P group, and 26.1% in MPG group. pCMV-Script/mito-MPG could be constructed and transfected into mitochondria of A549/DDP cells successfully, and inhibit proliferation and induce apoptosis of A549/DDP cells.

  5. Nitroxide-stimulated H2O2 decomposition by peroxidases and pseudoperoxidases.

    PubMed

    Mehlhorn, R J; Swanson, C E

    1992-01-01

    Nitroxide free radicals interact with Hb/metHb, Mb/metMb and with peroxidases/phenols to induce a catalase-like conversion of H2O2 to O2 (catalatic activity), without being substantially consumed in the process. The mechanism of this reaction is postulated to involve a one-electron oxidation of the nitroxide to the immonium oxene, which then reacts further to release oxygen and the nitroxide. An involvement of the immonium oxene in the reaction mechanism is consistent with ferryl heme reduction by nitroxides and a detection of the reduced nitroxide when the reaction mixture is supplemented with the two-electron reductant sodium borohydride. The nitroxide-induced catalatic activity is completely inhibited when the reaction mixture is supplemented with glutathione. Nitroxides suppress free radical formation by hydroperoxide-activated heme proteins, as inferred from their inhibition of the spin-trapping of glutathionyl radicals. H2O2 decomposition and a suppression of reactive free radical formation by heme proteins appears to be an antioxidant activity of nitroxides, which is distinct from their previously reported superoxide dismutating activity and which may be a factor in their protective action in models of cardiac reperfusion injury.

  6. Saturation factor of nitroxide radicals in liquid DNP by pulsed ELDOR experiments.

    PubMed

    Türke, Maria-Teresa; Bennati, Marina

    2011-03-07

    We propose the use of the pulse electron double resonance (ELDOR) method to determine the effective saturation factor of nitroxide radicals for dynamic nuclear polarization (DNP) experiments in liquids. The obtained values for the nitroxide radical TEMPONE-D,(15)N at different concentrations are rationalized in terms of spin relaxation and are shown to fulfil the Overhauser theory.

  7. High-Frequency Electron Paramagnetic Resonance Spectroscopy of Nitroxide-Functionalized Nanodiamonds in Aqueous Solution.

    PubMed

    Akiel, R D; Stepanov, V; Takahashi, S

    2016-06-21

    Nanodiamond (ND) is an attractive class of nanomaterial for fluorescent labeling, magnetic sensing of biological molecules, and targeted drug delivery. Many of those applications require tethering of target biological molecules on the ND surface. Even though many approaches have been developed to attach macromolecules to the ND surface, it remains challenging to characterize dynamics of tethered molecule. Here, we show high-frequency electron paramagnetic resonance (HF EPR) spectroscopy of nitroxide-functionalized NDs. Nitroxide radical is a commonly used spin label to investigate dynamics of biological molecules. In the investigation, we developed a sample holder to overcome water absorption of HF microwave. Then, we demonstrated HF EPR spectroscopy of nitroxide-functionalized NDs in aqueous solution and showed clear spectral distinction of ND and nitroxide EPR signals. Moreover, through EPR spectral analysis, we investigate dynamics of nitroxide radicals on the ND surface. The demonstration sheds light on the use of HF EPR spectroscopy to investigate biological molecule-functionalized nanoparticles.

  8. Nitroxides protect horseradish peroxidase from H2O2-induced inactivation and modulate its catalase-like activity.

    PubMed

    Samuni, Amram; Maimon, Eric; Goldstein, Sara

    2017-08-01

    Horseradish peroxidase (HRP) catalyzes H2O2 dismutation while undergoing heme inactivation. The mechanism underlying this process has not been fully elucidated. The effects of nitroxides, which protect metmyoglobin and methemoglobin against H2O2-induced inactivation, have been investigated. HRP reaction with H2O2 was studied by following H2O2 depletion, O2 evolution and heme spectral changes. Nitroxide concentration was followed by EPR spectroscopy, and its reactions with the oxidized heme species were studied using stopped-flow. Nitroxide protects HRP against H2O2-induced inactivation. The rate of H2O2 dismutation in the presence of nitroxide obeys zero-order kinetics and increases as [nitroxide] increases. Nitroxide acts catalytically since its oxidized form is readily reduced to the nitroxide mainly by H2O2. The nitroxide efficacy follows the order 2,2,6,6-tetramethyl-piperidine-N-oxyl (TPO)>4-OH-TPO>3-carbamoyl proxyl>4-oxo-TPO, which correlates with the order of the rate constants of nitroxide reactions with compounds I, II, and III. Nitroxide catalytically protects HRP against inactivation induced by H2O2 while modulating its catalase-like activity. The protective role of nitroxide at μM concentrations is attributed to its efficient oxidation by P940, which is the precursor of the inactivated form P670. Modeling the dismutation kinetics in the presence of nitroxide adequately fits the experimental data. In the absence of nitroxide the simulation fits the observed kinetics only if it does not include the formation of a Michaelis-Menten complex. Nitroxides catalytically protect heme proteins against inactivation induced by H2O2 revealing an additional role played by nitroxide antioxidants in vivo. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Isoindoline nitroxide-labeled porphyrins as potential fluorescence-suppressed spin probes.

    PubMed

    Liu, F; Zou, T J; Tan, Z L; Chen, S; Wu, Z H; Yan, G P; Zhang, Q; Liang, S C; Yang, J

    2017-02-07

    A series of isoindoline nitroxide-labeled porphyrins were synthesized by the reaction of 5-phenyldipyrromethane and 5-(4'-carboethoxy-methyleneoxyphenyl)dipyrromethane with 5-formyl-1,1,3,3-tetramethylisoindolin-2-yloxyl (FTMIO) using the Lindsey method. The corresponding water-soluble spin-labeled porphyrins were also prepared. Subsequently, these compounds were characterized and their in vitro properties were evaluated. The electrochemical assay demonstrated that these isoindoline nitroxide-labeled porphyrins had similar electrochemical and redox properties to 5-carboxy-1,1,3,3-tetramethylisoindolin-2-yloxyl (CTMIO). The electron paramagnetic resonance test showed that these porphyrins exhibited hyperfine splittings and characteristic spectra of CTMIO with typical nitroxide g-values and nitrogen isotropic hyperfine coupling constants. The in vitro cytotoxicity assay indicated that these porphyrins possessed low cytotoxicity to human renal tubular epithelial 293T cells (normal cells) and human hepatoma HepG2 cells (tumor cells). Fluorescence spectroscopy revealed that free base isoindoline nitroxide-labeled porphyrins exhibited fluorescence suppression characteristic of nitroxide-fluorophore systems. In vitro fluorescene imaging demonstrated that the reduced isoindoline nitroxide-labeled porphyrins eliminated fluorescence suppression and displayed strong red fluorescence imaging in HepG2 cells. Thus these isoindoline nitroxide-labeled porphyrins may be considered potentially as biological spin probes for fluorescence imaging and EPR spectroscopy.

  10. Mitochondria-targeted catalase reduces abnormal APP processing, amyloid β production and BACE1 in a mouse model of Alzheimer's disease: implications for neuroprotection and lifespan extension.

    PubMed

    Mao, Peizhong; Manczak, Maria; Calkins, Marcus J; Truong, Quang; Reddy, Tejaswini P; Reddy, Arubala P; Shirendeb, Ulziibat; Lo, Herng-Hsiang; Rabinovitch, Peter S; Reddy, P Hemachandra

    2012-07-01

    The purpose of this study was to investigate the protective effects of the mitochondria-targeted antioxidant catalase (MCAT) and lifespan extension in mice that express amyloid beta (Aβ). Using immunoblotting and immunostaining analyses, we measured the production of full-length amyloid precursor protein (APP), soluble APPα, C-terminal fragments CTF99 and CTF83, monomeric and oligomeric Aβ, Aβ deposits and beta site amyloid precursor protein cleaving enzyme 1 (BACE1), in different stages of disease progression in MCAT/AβPP and AβPP mice. Using quantitative reverse transcriptase polymerase chain reaction and immunostaining analyses, we studied the expression of catalase, BACE1, the Alzheimer's disease (AD) markers, synaptophysin, APP, neprilysin, insulin-degrading enzyme and transthyretin in MCAT, AβPP, MCAT/AβPP and wild-type (WT) mice. Using the high pressure liquid chromatography analysis of 8-hydroxy-2-deoxyguanosine, we measured oxidative DNA damage in the cerebral cortical tissues from MCAT, AβPP, MCAT/AβPP and WT mice. We found that the AβPP transgenic mice that carried the human MCAT gene lived 5 months longer than did the AβPP mice. We also found that the overexpression of MCAT in the brain sections from the MCAT/AβPP transgenic mice significantly correlated with a reduction in the levels of full-length APP, CTF99, BACE1, Aβ levels (40 and 42), Aβ deposits and oxidative DNA damage relative to the brain sections from the AβPP mice. Interestingly, we found significantly increased levels of soluble APPα and CTF83 in the MCAT/AβPP mice, relative to the AβPP mice. These data provide direct evidence that oxidative stress plays a primary role in AD etiopathology and that in MCAT mice express Aβ, MCAT prevents abnormal APP processing, reduces Aβ levels and enhances Aβ-degrading enzymes in mice at different ages, corresponding to different stages of disease progression. These findings indicate that mitochondria-targeted molecules may be an

  11. Chemistry and Antihypertensive Effects of Tempol and Other Nitroxides

    PubMed Central

    WILCOX, CHRISTOPHER S.; PEARLMAN, ADAM

    2009-01-01

    Nitroxides can undergo one- or two-electron reduction reactions to hydroxylamines or oxammonium cations, respectively, which themselves are interconvertible, thereby providing redox metabolic actions. 4-Hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (tempol) is the most extensively studied nitroxide. It is a cell membrane-permeable amphilite that dismutates superoxide catalytically, facilitates hydrogen peroxide metabolism by catalase-like actions, and limits formation of toxic hydroxyl radicals produced by Fenton reactions. It is broadly effective in detoxifying these reactive oxygen species in cell and animal studies. When administered intravenously to hypertensive rodent models, tempol caused rapid and reversible dose-dependent reductions in blood pressure in 22 of 26 studies. This was accompanied by vasodilation, increased nitric oxide activity, reduced sympathetic nervous system activity at central and peripheral sites, and enhanced potassium channel conductance in blood vessels and neurons. When administered orally or by infusion over days or weeks to hypertensive rodent models, it reduced blood pressure in 59 of 68 studies. This was accompanied by correction of salt sensitivity and endothelial dysfunction and reduced agonist-evoked oxidative stress and contractility of blood vessels, reduced renal vascular resistance, and increased renal tissue oxygen tension. Thus, tempol is broadly effective in reducing blood pressure, whether given by acute intravenous injection or by prolonged administration, in a wide range of rodent models of hypertension. PMID:19112152

  12. The mitochondria targeted antioxidant MitoQ protects against fluoroquinolone-induced oxidative stress and mitochondrial membrane damage in human Achilles tendon cells.

    PubMed

    Lowes, Damon A; Wallace, Carol; Murphy, Michael P; Webster, Nigel R; Galley, Helen F

    2009-04-01

    Tendinitis and tendon rupture during treatment with fluoroquinolone antibiotics is thought to be mediated via oxidative stress. This study investigated whether ciprofloxacin and moxifloxacin cause oxidative stress and mitochondrial damage in cultured normal human Achilles' tendon cells and whether an antioxidant targeted to mitochondria (MitoQ) would protect against such damage better than a non-mitochondria targeted antioxidant. Human tendon cells from normal Achilles' tendons were exposed to 0-0.3 mM antibiotic for 24 h and 7 days in the presence of 1 microM MitoQ or an untargeted form, idebenone. Both moxifloxacin and ciprofloxacin resulted in up to a 3-fold increase in the rate of oxidation of dichlorodihydrofluorescein, a marker of general oxidative stress in tenocytes (p<0.0001) and loss of mitochondrial membrane permeability (p<0.001). In cells treated with MitoQ the oxidative stress was less and mitochondrial membrane potential was maintained. Mitochondrial damage to tenocytes during fluoroquinolone treatment may be involved in tendinitis and tendon rupture.

  13. A novel class of mitochondria-targeted soft electrophiles modifies mitochondrial proteins and inhibits mitochondrial metabolism in breast cancer cells through redox mechanisms.

    PubMed

    Vayalil, Praveen K; Oh, Joo-Yeun; Zhou, Fen; Diers, Anne R; Smith, M Ryan; Golzarian, Hafez; Oliver, Patsy G; Smith, Robin A J; Murphy, Michael P; Velu, Sadanandan E; Landar, Aimee

    2015-01-01

    Despite advances in screening and treatment over the past several years, breast cancer remains a leading cause of cancer-related death among women in the United States. A major goal in breast cancer treatment is to develop safe and clinically useful therapeutic agents that will prevent the recurrence of breast cancers after front-line therapeutics have failed. Ideally, these agents would have relatively low toxicity against normal cells, and will specifically inhibit the growth and proliferation of cancer cells. Our group and others have previously demonstrated that breast cancer cells exhibit increased mitochondrial oxygen consumption compared with non-tumorigenic breast epithelial cells. This suggests that it may be possible to deliver redox active compounds to the mitochondria to selectively inhibit cancer cell metabolism. To demonstrate proof-of-principle, a series of mitochondria-targeted soft electrophiles (MTSEs) has been designed which selectively accumulate within the mitochondria of highly energetic breast cancer cells and modify mitochondrial proteins. A prototype MTSE, IBTP, significantly inhibits mitochondrial oxidative phosphorylation, resulting in decreased breast cancer cell proliferation, cell attachment, and migration in vitro. These results suggest MTSEs may represent a novel class of anti-cancer agents that prevent cancer cell growth by modification of specific mitochondrial proteins.

  14. Chloroplast- or Mitochondria-Targeted DEAD-Box RNA Helicases Play Essential Roles in Organellar RNA Metabolism and Abiotic Stress Responses

    PubMed Central

    Nawaz, Ghazala; Kang, Hunseung

    2017-01-01

    The yields and productivity of crops are greatly diminished by various abiotic stresses, including drought, cold, heat, and high salinity. Chloroplasts and mitochondria are cellular organelles that can sense diverse environmental stimuli and alter gene expression to cope with adverse environmental stresses. Organellar gene expression is mainly regulated at posttranscriptional levels, including RNA processing, intron splicing, RNA editing, RNA turnover, and translational control, during which a variety of nucleus-encoded RNA-binding proteins (RBPs) are targeted to chloroplasts or mitochondria where they play essential roles in organellar RNA metabolism. DEAD-box RNA helicases (RHs) are enzymes that can alter RNA structures and affect RNA metabolism in all living organisms. Although a number of DEAD-box RHs have been found to play important roles in RNA metabolism in the nucleus and cytoplasm, our understanding on the roles of DEAD-box RHs in the regulation of RNA metabolism in chloroplasts and mitochondria is only at the beginning. Considering that organellar RNA metabolism and gene expression are tightly regulated by anterograde signaling from the nucleus, it is imperative to determine the functions of nucleus-encoded organellar RBPs. In this review, we summarize the emerging roles of nucleus-encoded chloroplast- or mitochondria-targeted DEAD-box RHs in organellar RNA metabolism and plant response to diverse abiotic stresses. PMID:28596782

  15. Chloroplast- or Mitochondria-Targeted DEAD-Box RNA Helicases Play Essential Roles in Organellar RNA Metabolism and Abiotic Stress Responses.

    PubMed

    Nawaz, Ghazala; Kang, Hunseung

    2017-01-01

    The yields and productivity of crops are greatly diminished by various abiotic stresses, including drought, cold, heat, and high salinity. Chloroplasts and mitochondria are cellular organelles that can sense diverse environmental stimuli and alter gene expression to cope with adverse environmental stresses. Organellar gene expression is mainly regulated at posttranscriptional levels, including RNA processing, intron splicing, RNA editing, RNA turnover, and translational control, during which a variety of nucleus-encoded RNA-binding proteins (RBPs) are targeted to chloroplasts or mitochondria where they play essential roles in organellar RNA metabolism. DEAD-box RNA helicases (RHs) are enzymes that can alter RNA structures and affect RNA metabolism in all living organisms. Although a number of DEAD-box RHs have been found to play important roles in RNA metabolism in the nucleus and cytoplasm, our understanding on the roles of DEAD-box RHs in the regulation of RNA metabolism in chloroplasts and mitochondria is only at the beginning. Considering that organellar RNA metabolism and gene expression are tightly regulated by anterograde signaling from the nucleus, it is imperative to determine the functions of nucleus-encoded organellar RBPs. In this review, we summarize the emerging roles of nucleus-encoded chloroplast- or mitochondria-targeted DEAD-box RHs in organellar RNA metabolism and plant response to diverse abiotic stresses.

  16. The mitochondria-targeted antioxidant MitoQ extends lifespan and improves healthspan of a transgenic Caenorhabditis elegans model of Alzheimer disease.

    PubMed

    Ng, Li Fang; Gruber, Jan; Cheah, Irwin K; Goo, Chong Kit; Cheong, Wei Fun; Shui, Guanghou; Sit, Kim Ping; Wenk, Markus R; Halliwell, Barry

    2014-06-01

    β-Amyloid (Aβ)-induced toxicity and oxidative stress have been postulated to play critical roles in the pathogenic mechanism of Alzheimer disease (AD). We investigated the in vivo ability of a mitochondria-targeted antioxidant, MitoQ, to protect against Aβ-induced toxicity and oxidative stress in a Caenorhabditis elegans model overexpressing human Aβ. Impairment of electron transport chain (ETC) enzymatic activity and mitochondrial dysfunction are early features of AD. We show that MitoQ extends lifespan, delays Aβ-induced paralysis, ameliorates depletion of the mitochondrial lipid cardiolipin, and protects complexes IV and I of the ETC. Despite its protective effects on lifespan, healthspan, and ETC function, we find that MitoQ does not reduce DCFDA fluorescence, protein carbonyl levels or modulate steadystate ATP levels or oxygen consumption rate. Moreover, MitoQ does not attenuate mitochondrial DNA (mtDNA) oxidative damage. In agreement with its design, the protective effects of MitoQ appear to be targeted specifically to the mitochondrial membrane and our findings suggest that MitoQ may have therapeutic potential for Aβ- and oxidative stress-associated neurodegenerative disorders, particularly AD. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  17. A Novel Class of Mitochondria-Targeted Soft Electrophiles Modifies Mitochondrial Proteins and Inhibits Mitochondrial Metabolism in Breast Cancer Cells through Redox Mechanisms

    PubMed Central

    Vayalil, Praveen K.; Oh, Joo-Yeun; Zhou, Fen; Diers, Anne R.; Smith, M. Ryan; Golzarian, Hafez; Oliver, Patsy G.; Smith, Robin A. J.; Murphy, Michael P.; Velu, Sadanandan E.; Landar, Aimee

    2015-01-01

    Despite advances in screening and treatment over the past several years, breast cancer remains a leading cause of cancer-related death among women in the United States. A major goal in breast cancer treatment is to develop safe and clinically useful therapeutic agents that will prevent the recurrence of breast cancers after front-line therapeutics have failed. Ideally, these agents would have relatively low toxicity against normal cells, and will specifically inhibit the growth and proliferation of cancer cells. Our group and others have previously demonstrated that breast cancer cells exhibit increased mitochondrial oxygen consumption compared with non-tumorigenic breast epithelial cells. This suggests that it may be possible to deliver redox active compounds to the mitochondria to selectively inhibit cancer cell metabolism. To demonstrate proof-of-principle, a series of mitochondria-targeted soft electrophiles (MTSEs) has been designed which selectively accumulate within the mitochondria of highly energetic breast cancer cells and modify mitochondrial proteins. A prototype MTSE, IBTP, significantly inhibits mitochondrial oxidative phosphorylation, resulting in decreased breast cancer cell proliferation, cell attachment, and migration in vitro. These results suggest MTSEs may represent a novel class of anti-cancer agents that prevent cancer cell growth by modification of specific mitochondrial proteins. PMID:25785718

  18. Synthesis and Evaluation of Ciprofloxacin-Nitroxide Conjugates as Anti-Biofilm Agents.

    PubMed

    Verderosa, Anthony D; Mansour, Sarah C; de la Fuente-Núñez, César; Hancock, Robert E W; Fairfull-Smith, Kathryn E

    2016-06-27

    As bacterial biofilms are often refractory to conventional antimicrobials, the need for alternative and/or novel strategies for the treatment of biofilm related infections has become of paramount importance. Herein, we report the synthesis of novel hybrid molecules comprised of two different hindered nitroxides linked to the piperazinyl secondary amine of ciprofloxacin via a tertiary amine linker achieved utilising reductive amination. The corresponding methoxyamine derivatives were prepared alongside their radical-containing counterparts as controls. Subsequent biological evaluation of the hybrid compounds on preformed P. aeruginosa flow cell biofilms divulged significant dispersal and eradication abilities for ciprofloxacin-nitroxide hybrid compound 10 (up to 95% eradication of mature biofilms at 40 μM). Importantly, these hybrids represent the first dual-action antimicrobial-nitroxide agents, which harness the dispersal properties of the nitroxide moiety to circumvent the well-known resistance of biofilms to treatment with antimicrobial agents.

  19. Silica materials with wall-embedded nitroxides provide efficient polarization matrices for dynamic nuclear polarization NMR.

    PubMed

    Besson, Eric; Ziarelli, Fabio; Bloch, Emily; Gerbaud, Guillaume; Queyroy, Séverine; Viel, Stéphane; Gastaldi, Stéphane

    2016-04-25

    Hybrid mesoporous silica materials with wall-embedded nitroxides are shown to efficiently polarize impregnated substrates in high-field dynamic nuclear polarization magic-angle spinning solid-state NMR experiments.

  20. Nitroxide free radicals protect macular carotenoids against chemical destruction (bleaching) during lipid peroxidation.

    PubMed

    Zareba, M; Widomska, J; Burke, J M; Subczynski, W K

    2016-12-01

    Macular xanthophylls (MXs) lutein and zeaxanthin are dietary carotenoids that are selectively concentrated in the human eye retina, where they are thought to protect against age-related macular degeneration (AMD) by multiple mechanisms, including filtration of phototoxic blue light and quenching of singlet oxygen and triplet states of photosensitizers. These physical protective mechanisms require that MXs be in their intact structure. Here, we investigated the protection of the intact structure of zeaxanthin incorporated into model membranes subjected to oxidative modification by water- and/or membrane-soluble small nitroxide free radicals. Model membranes were formed from saturated, monounsaturated, and polyunsaturated phosphatidylcholines (PCs). Oxidative modification involved autoxidation, iron-mediated, and singlet oxygen-mediated lipid peroxidation. The extent of chemical destruction (bleaching) of zeaxanthin was evaluated from its absorption spectra and compared with the extent of lipid peroxidation evaluated using the thiobarbituric acid assay. Nitroxide free radicals with different polarity (membrane/water partition coefficients) were used. The extent of zeaxanthin bleaching increased with membrane unsaturation and correlated with the rate of PC oxidation. Protection of the intact structure of zeaxanthin by membrane-soluble nitroxides was much stronger than that by water-soluble nitroxides. The combination of zeaxanthin and lipid-soluble nitroxides exerted strong synergistic protection against singlet oxygen-induced lipid peroxidation. The synergistic effect may be explained in terms of protection of the intact zeaxanthin structure by effective scavenging of free radicals by nitroxides, therefore allowing zeaxanthin to quench the primary oxidant, singlet oxygen, effectively by the physical protective mechanism. The redox state of nitroxides was monitored using electron paramagnetic resonance spectroscopy. Both nitroxide free radicals and their reduced form

  1. Photoinduced electron transfer from dialkyl nitroxides to halogenated solvents

    SciTech Connect

    Chateauneuf, J. ); Lusztyk, J.; Ingold, K.U. )

    1990-02-02

    Laser flash photolysis (LFP) at wavelengths within the charge-transfer absorption present in CCl{sub 4} solutions of 2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPO) yields the oxoammonium chloride of TEMPO, 1 ({lambda}{sub max} = 460 nm), and the trichloromethyl radical in an essentially instantaneous ({le}18 ps) process. The primary photochemical event is an electron transfer from TEMPO to CCl{sub 4}, and this is followed by immediate decomposition of the CCl{sub 4}{sup {sm bullet}{minus}} radical anion to Cl{sup {minus}} and Cl{sub 3}C{sup {sm bullet}}. An independent synthesis of 1 confirmed that the absorption attributed to this species has been correctly assigned. The formation of Cl{sub 3}C{sup {sm bullet}} was inferred by its trapping by molecular oxygen. LFP of TEMPO in other halogenated solvents and of other nitroxides in halogenated solvents has confirmed the generality of these photoreactions.

  2. Magnetic phenomena in nonrigid metal-nitroxide systems

    NASA Astrophysics Data System (ADS)

    Ovcharenko, V.; Fursova, E.; Fokin, S.; Romanenko, G.; Ikorskii, V.

    2004-04-01

    For NiL{2} heterospin bischelates, stereochemical nonrigidity in solution has been found, which leads to solids with varying structure and composition. While investigating the products of Cu(hfac){2} interaction with spin-labeled pyrazole 4,4,5,5-tetramethyl-2-(1-methyl-1H-pyrazol-4-yl)-imidazoline-3-oxide-1-oxyl we have isolated a family of heterospin compounds differing in the structure and composition in the solid state. In synthetic systems, these compounds often co-crystallize and must be separated mechanically. It is also shown that minor variations in the structure of solid heterospin complexes substantially change the magnetic properties of the compounds. Key words. Nitroxides - metal complexes structure - magnetic properties.

  3. Antioxidant effects of water- and lipid-soluble nitroxide radicals in liposomes.

    PubMed

    Cimato, Alejandra N; Piehl, Lidia L; Facorro, Graciela B; Torti, Horacio B; Hager, Alfredo A

    2004-12-15

    Liposomes are today useful tools in different fields of science and technology. A lack of stability due to lipid peroxidation is the main problem in the extension of the use of these formulations. Recent investigative works have reported the protective effects of stable nitroxide radicals against oxidative processes in different media and under different stress conditions. Our group has focused its attention on the natural aging of liposomes and the protection provided by the water- and lipid-soluble nitroxide radicals 2,2,6,6-tetramethylpiperdine-1-oxyl (TEMPO) and doxylstearic acids (5-DSA, 12-DSA, and 16-DSA), respectively. Unilamellar liposomes were incubated under air atmosphere at 37 degrees C, both in the absence and in the presence of these radicals. Conjugated dienes, lipid hydroperoxides, TBARS, membrane fluidity, and nitroxide ESR signal intensity were followed as a function of time. Our results demonstrated that doxylstearic acids were more efficient than TEMPO in retarding lipid peroxidation at all the concentrations tested. The inhibition percentages, depending on the total nitroxide concentration, were not proportional to the lipid-water partition coefficient. Furthermore, time-course ESR signals showed a slower decrease for doxylstearic acids than for TEMPO. No significant differences were found among 5-DSA, 12-DSA, and 16-DSA. We concluded that the nitroxide radical efficiency as antioxidant directly depends on both nitroxide concentration and lipophilicity.

  4. A comparison between nitroxide and hydrazyl free radicals in selective alcohols oxidation

    NASA Astrophysics Data System (ADS)

    Shakir, Ahmed J.; Madalan, Augustin M.; Ionita, Gabriela; Lupu, Stelian; Lete, Cecilia; Ionita, Petre

    2017-06-01

    Literature data are abundant in oxidations catalyzed by nitroxides free radicals, while hydrazyls free radicals are very seldom encountered. In this work we made a comparison between some nitroxides and hydrazyl free radicals towards the ability to selectively oxidize activated alcohols to the corresponding carbonyl derivatives. As nitroxides we used 2,2,6,6-tetramethylpirrolidin-N-oxide (TEMPO) and phtalimide-N-oxyl (PINO) free radicals, while as hydrazyls were used 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-p-nitrophenyl-1-picrylhydrazyl (DN-DPPH). The differences in the physico-chemical properties between nitroxides and hydrazyls were evaluated by UV-vis, EPR and cyclic voltammetry. For DN-DPPH the X-ray crystal structure was resolved. It has been shown that a nitroxide radical is a far better catalyst in such aerobic oxidations comparatively with a hydrazyl one. The explanation of these results consists firstly in a different mechanism involved in the oxidation procedures and secondly in the variation of the oxidation potentials of nitroxides comparatively with hydrazyls.

  5. Syntheses, structures and magnetic properties of four-spin Mn-Imino nitroxide radical complexes

    NASA Astrophysics Data System (ADS)

    Lv, Xue-Hui; Yang, Shuai-Liang; Li, Yuan-Xia; Zhang, Chen-Xi; Wang, Qing-Lun

    2017-04-01

    Based on the nitroxide radicals, four-spin complexes [Mn(hfac)2(IMpPhCl)]2·NITpPhCl (1) and [Mn(hfac)2(IMmPhCl)]2·NITmPhCl (2) (IMpPhCl = 2-(4'-chlorophenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl, IMmPhCl = 2-(3'-chlorophenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl, hfac = hexafluoroacetylacetonate) have been synthesized and characterized by single-crystal X-ray diffraction. The X-ray crystal structure analyses show that the structures of the two compounds are similar and the imino nitroxide radical ligand acts as a bridge ligand linking two Mn(II) ions through the oxygen atom of the N-O group to form a four-spin system. Two kinds of nitroxide radicals: nitronyl nitroxide and imino nitroxide radicals coexist in the cyclic Mn(II) complexes. The magnetic studies show that there exists an antiferromagnetic interaction between Mn(II) ions and the imino nitroxide radical ligands, which is explained by spin polarization mechanism. The antiferromagnetic interaction of Mn-Rad in complex 2 (J1 = -9.36 cm-1) is stronger than that in complex 1 (J1 = -9.19 cm-1), which is consistent with crystal structure of complexes (The bond length of the shortest Mn-O in complex 2 (2.1625 Å) is smaller than complex 1 (2.1898 Å)).

  6. Partitioning of nitroxides in dispersed systems investigated by ultrafiltration, EPR and NMR spectroscopy.

    PubMed

    Krudopp, Heimke; Sönnichsen, Frank D; Steffen-Heins, Anja

    2015-08-15

    The partitioning behavior of paramagnetic nitroxides in dispersed systems can be determined by deconvolution of electron paramagnetic resonance (EPR) spectra giving equivalent results with the validated methods of ultrafiltration techniques (UF) and pulsed-field gradient nuclear magnetic resonance spectroscopy (PFG-NMR). The partitioning behavior of nitroxides with increasing lipophilicity was investigated in anionic, cationic and nonionic micellar systems and 10 wt% o/w emulsions. Apart from EPR spectra deconvolution, the PFG-NMR was used in micellar solutions as a non-destructive approach, while UF based on separation of very small volume of the aqueous phase. As a function of their substituent and lipophilicity, the proportions of nitroxides that were solubilized in the micellar or emulsion interface increased with increasing nitroxide lipophilicity for all emulsifier used. Comparing the different approaches, EPR deconvolution and UF revealed comparable nitroxide proportions that were solubilized in the interfaces. Those proportions were higher than found with PFG-NMR. For PFG-NMR self-diffusion experiments the reduced nitroxides were used revealing a high dynamic of hydroxylamines and emulsifiers. Deconvolution of EPR spectra turned out to be the preferred method for measuring the partitioning behavior of paramagnetic molecules as it enables distinguishing between several populations at their individual solubilization sites. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. An ESR study of the nitroxide radical of pentastarch-conjugated deferoxamine

    SciTech Connect

    Pieper, G.M.; Gross, G.J.; Kalyanaraman, B. )

    1990-01-01

    At higher concentrations, deferoxamine (DFO) reacts with hydroxyl radicals to produce a stable nitroxide free radical. Formation and decay of this nitroxide radical was investigated and compared with a novel modified pentastarch conjugate of DFO (MPS-DFO). Photolytic generation of hydroxyl radicals from H2O2 in the presence of free DFO produced a nitroxide radical with coupling constants of aN = 8.0 G and aH = 6.5 G. Under the same experimental conditions, equimolar concentrations of MPS-DFO produced an ESR signal of reduced intensity while iron-saturated MPS-DFO produced no signal. Incubation of free DFO with pentastarch (i.e., without conjugation) greatly decreased the intensity of the nitroxide radical signal. Using a spin-trapping technique with 5,5-dimethyl-1-pyrroline N-oxide (DMPO), the pentastarch vehicle was shown to inhibit the DMPO-OH adduct formation. The decay of the DFO nitroxide radical decayed with a second-order rate constant while that of MPS-DFO decayed with a first-order rate constant. Thus, a novel derivative of DFO may provide some additional benefit in limiting DFO nitroxide radical formation and might explain the reported reduced in vivo toxicity of MPS-DFO relative to free DFO.

  8. Integrated experimental and computational spectroscopy study on the protonation of the α-nitronyl nitroxide radical unit.

    PubMed

    Pütz, Anna-Maria; Schatzschneider, Ulrich; Rentschler, Eva

    2012-02-07

    The stability of the α-nitronyl nitroxide radical unit under acidic conditions is investigated by an integrated experimental UV/Vis spectroscopy and TDDFT study. In the field of molecular magnetism, α-nitronyl nitroxide radicals are important as purely organic spin carriers due to their synthetic versatility. Here, the existence of an intermediate of the protonated α-nitronyl nitroxide radical unit is demonstrated for the first time and a proposed disproportionation reaction is confirmed.

  9. Development of a mitochondria-based centrifugal ultrafiltration/liquid chromatography/mass spectrometry method for screening mitochondria-targeted bioactive constituents from complex matrixes: Herbal medicines as a case study.

    PubMed

    Yang, Xing-Xin; Xu, Feng; Wang, Dan; Yang, Zhi-Wei; Tan, Huan-Ran; Shang, Ming-Ying; Wang, Xuan; Cai, Shao-Qing

    2015-09-25

    Mitochondria are an important intracellular pharmacological target because damage to this organelle results in a variety of human disorders and because mitochondria are involved in complex processes such as energy generation, apoptosis and lipid metabolism. To expedite the search for natural bioactive compounds targeting mitochondria, we initially developed an efficient mitochondria-based screening method by combining centrifugal ultrafiltration (CU) with liquid chromatography/mass spectrometry (LC/MS), which is called screening method for mitochondria-targeted bioactive constituents (SM-MBC) and is compatible with the search of mitochondria-targeted compounds from complex matrixes such as herbal medicines extracts. Functionally active, structurally intact and pure mitochondria were obtained from rat myocardium using an optimized protocol for mitochondrial isolation comprising organelle release followed by differential and Nycodenz density gradient centrifugation. After evaluating the reliability of the method using thiabendazole (TZ), rotenone (RN), amiodarone (AR) and trimetazidine (TD) as positive controls, this method was successfully applied to screen bioactive constituents from extracts of Polygoni Cuspidati Rhizoma et Radix (PCRR) and Scutellariae Radix (SR). Nineteen active compounds were detected and identified by LC/MS, of which 17 were new mitochondria-targeted compounds. The activity of 9 of the 19 hit compounds was confirmed by in vitro pharmacological trials. These results demonstrate that SM-MBC can be used for the efficient screening of mitochondria-targeted constituents in complex preparations used to treat mitochondrial disorders, such as PCRR and SR. The results may be meaningful for an in-depth understanding of drug mechanism of action and drug discovery from medicinal herbs.

  10. The Mitochondria-Targeted Antioxidant SkQ1 Downregulates Aryl Hydrocarbon Receptor-Dependent Genes in the Retina of OXYS Rats with AMD-Like Retinopathy

    PubMed Central

    Perepechaeva, M. L.; Grishanova, A. Yu.; Rudnitskaya, E. A.; Kolosova, N. G.

    2014-01-01

    The mitochondria-targeted antioxidant SkQ1 is a novel drug thought to retard development of age-related diseases. It has been shown that SkQ1 reduces clinical signs of retinopathy in senescence-accelerated OXYS rats, which are a known animal model of human age-related macular degeneration (AMD). The aim of this work was to test whether SkQ1 affects transcriptional activity of AhR (aryl hydrocarbon receptor) and Nrf2 (nuclear factor erythroid 2-related factor 2), which are considered as AMD-associated genes in the retina of OXYS and Wistar rats. Our results showed that only AhR and AhR-dependent genes were sensitive to SkQ1. Dietary supplementation with SkQ1 decreased the AhR mRNA level in both OXYS and Wistar rats. At baseline, the retinal Cyp1a1 mRNA level was lower in OXYS rats. SkQ1 supplementation decreased the Cyp1a1 mRNA level in Wistar rats, but this level remained unchanged in OXYS rats. Baseline Cyp1a2 and Cyp1b1 mRNA expression was stronger in OXYS than in Wistar rats. In the OXYS strain, Cyp1a2 and Cyp1b1 mRNA levels decreased as a result of SkQ1 supplementation. These data suggest that the Cyp1a2 and Cyp1b1 enzymes are involved in the pathogenesis of AMD-like retinopathy of OXYS rats and are possible therapeutic targets of SkQ1. PMID:25132985

  11. Respective contribution of mitochondrial superoxide and pH to mitochondria-targeted circularly permuted yellow fluorescent protein (mt-cpYFP) flash activity.

    PubMed

    Wei-LaPierre, Lan; Gong, Guohua; Gerstner, Brent J; Ducreux, Sylvie; Yule, David I; Pouvreau, Sandrine; Wang, Xianhua; Sheu, Shey-Shing; Cheng, Heping; Dirksen, Robert T; Wang, Wang

    2013-04-12

    Superoxide flashes are transient bursts of superoxide production within the mitochondrial matrix that are detected using the superoxide-sensitive biosensor, mitochondria-targeted circularly permuted YFP (mt-cpYFP). However, due to the pH sensitivity of mt-cpYFP, flashes were suggested to reflect transient events of mitochondrial alkalinization. Here, we simultaneously monitored flashes with mt-cpYFP and mitochondrial pH with carboxy-SNARF-1. In intact cardiac myocytes and purified skeletal muscle mitochondria, robust mt-cpYFP flashes were accompanied by only a modest increase in SNARF-1 ratio (corresponding to a pH increase of <0.1), indicating that matrix alkalinization is minimal during an mt-cpYFP flash. Individual flashes were also accompanied by stepwise increases of MitoSOX signal and decreases of NADH autofluorescence, supporting the superoxide origin of mt-cpYFP flashes. Transient matrix alkalinization induced by NH4Cl only minimally influenced flash frequency and failed to alter flash amplitude. However, matrix acidification modulated superoxide flash frequency in a bimodal manner. Low concentrations of nigericin (< 100 nM) that resulted in a mild dissipation of the mitochondrial pH gradient increased flash frequency, whereas a maximal concentration of nigericin (5 μm) collapsed the pH gradient and abolished flash activity. These results indicate that mt-cpYFP flash events reflect a burst in electron transport chain-dependent superoxide production that is coincident with a modest increase in matrix pH. Furthermore, flash activity depends strongly on a combination of mitochondrial oxidation and pH gradient.

  12. MitoQ, a mitochondria-targeted antioxidant, delays disease progression and alleviates pathogenesis in an experimental autoimmune encephalomyelitis mouse model of multiple sclerosis.

    PubMed

    Mao, Peizhong; Manczak, Maria; Shirendeb, Ulziibat P; Reddy, P Hemachandra

    2013-12-01

    Oxidative stress and mitochondrial dysfunction are involved in the progression and pathogenesis of multiple sclerosis (MS). MitoQ is a mitochondria-targeted antioxidant that has a neuroprotective role in several mitochondrial and neurodegenerative diseases, including Alzheimer's disease and Parkinson's disease. Here we sought to determine the possible effects of a systematic administration of MitoQ as a therapy, using an experimental autoimmune encephalomyelitis (EAE) mouse model. We studied the beneficial effects of MitoQ in EAE mice that mimic MS like symptoms by treating EAE mice with MitoQ and pretreated C57BL6 mice with MitoQ plus EAE induction. We found that pretreatment and treatment of EAE mice with MitoQ reduced neurological disabilities associated with EAE. We also found that both pretreatment and treatment of the EAE mice with MitoQ significantly suppressed inflammatory markers of EAE, including the inhibition of inflammatory cytokines and chemokines. MitoQ treatments reduced neuronal cell loss in the spinal cord, a factor underlying motor disability in EAE mice. The neuroprotective role of MitoQ was confirmed by a neuron-glia co-culture system designed to mimic the mechanism of MS and EAE in vitro. We found that axonal inflammation and oxidative stress are associated with impaired behavioral functions in the EAE mouse model and that treatment with MitoQ can exert protective effects on neurons and reduce axonal inflammation and oxidative stress. These protective effects are likely via multiple mechanisms, including the attenuation of the robust immune response. These results suggest that MitoQ may be a new candidate for the treatment of MS. © 2013.

  13. Mitochondria-targeted DsRed2 protein expression during the early stage of bovine somatic cell nuclear transfer embryo development.

    PubMed

    Park, Hyo-Jin; Min, Sung-Hun; Choi, Hoonsung; Park, Junghyung; Kim, Sun-Uk; Lee, Seunghoon; Lee, Sang-Rae; Kong, Il-Keun; Chang, Kyu-Tae; Koo, Deog-Bon; Lee, Dong-Seok

    2016-09-01

    Somatic cell nuclear transfer (SCNT) has been widely used as an efficient tool in biomedical research for the generation of transgenic animals from somatic cells with genetic modifications. Although remarkable advances in SCNT techniques have been reported in a variety of mammals, the cloning efficiency in domestic animals is still low due to the developmental defects of SCNT embryos. In particular, recent evidence has revealed that mitochondrial dysfunction is detected during the early development of SCNT embryos. However, there have been relatively few or no studies regarding the development of a system for evaluating mitochondrial behavior or dynamics. For the first time, in mitochondria of bovine SCNT embryos, we developed a method for the visualization of mitochondria and expression of fluorescence proteins. To express red fluorescence in mitochondria of cloned embryos, bovine ear skin fibroblasts, nuclear donor, were stably transfected with a vector carrying mitochondria-targeting DsRed2 gene tagged with V5 epitope (mito-DsRed2-V5 tag) using lentivirus-mediated gene transfer because of its ability to integrate in the cell genome and the potential for long-term transgene expression in the transduced cells and their dividing cells. From western blotting analysis of V5 tag protein using mitochondrial fraction and confocal microscopy of red fluorescence using SCNT embryos, we found that the mitochondrial expression of the mito-DsRed2 protein was detected until the blastocyst stage. In addition, according to image analysis, it may be suggested possible use of the system for visualization of mitochondrial localization and evaluation of mitochondrial behaviors or dynamics in early development of bovine SCNT embryos.

  14. The novel mitochondria-targeted hydrogen sulfide (H2S) donors AP123 and AP39 protect against hyperglycemic injury in microvascular endothelial cells in vitro.

    PubMed

    Gerő, Domokos; Torregrossa, Roberta; Perry, Alexis; Waters, Alicia; Le-Trionnaire, Sophie; Whatmore, Jacqueline L; Wood, Mark; Whiteman, Matthew

    2016-11-01

    The development of diabetic vascular complications is initiated, at least in part, by mitochondrial reactive oxygen species (ROS) production in endothelial cells. Hyperglycemia induces superoxide production in the mitochondria and initiates changes in the mitochondrial membrane potential that leads to mitochondrial dysfunction. Hydrogen sulfide (H2S) supplementation has been shown to reduce the mitochondrial oxidant production and shows efficacy against diabetic vascular damage in vivo. However, the half-life of H2S is very short and it is not specific for the mitochondria. We have therefore evaluated two novel mitochondria-targeted anethole dithiolethione and hydroxythiobenzamide H2S donors (AP39 and AP123 respectively) at preventing hyperglycemia-induced oxidative stress and metabolic changes in microvascular endothelial cells in vitro. Hyperglycemia (HG) induced significant increase in the activity of the citric acid cycle and led to elevated mitochondrial membrane potential. Mitochondrial oxidant production was increased and the mitochondrial electron transport decreased in hyperglycemic cells. AP39 and AP123 (30-300nM) decreased HG-induced hyperpolarisation of the mitochondrial membrane and inhibited the mitochondrial oxidant production. Both H2S donors (30-300nM) increased the electron transport at respiratory complex III and improved the cellular metabolism. Targeting H2S to mitochondria retained the cytoprotective effect of H2S against glucose-induced damage in endothelial cells suggesting that the molecular target of H2S action is within the mitochondria. Mitochondrial targeting of H2S also induced >1000-fold increase in the potency of H2S against hyperglycemia-induced injury. The high potency and long-lasting effect elicited by these H2S donors strongly suggests that these compounds could be useful against diabetic vascular complications. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  15. NOVEL MITOCHONDRIA-TARGETED ANTIOXIDANT PEPTIDE AMELIORATES BURN-INDUCED APOPTOSIS AND ENDOPLASMIC RETICULUM STRESS IN THE SKELETAL MUSCLE OF MICE

    PubMed Central

    Lee, Hyung-yul; Kaneki, Masao; Andreas, Jonathan; Tompkins, Ronald G.; Martyn, J.A. Jeevendra

    2011-01-01

    This study tested the hypothesis that a novel mitochondria-targeted SS-31 peptide attenuates the burn injury-induced apoptosis and endoplasmic reticulum (ER) stress and improves insulin sensitivity in the skeletal muscle. Following 30% total body surface area burn or sham-burn, mice were injected daily with SS-31 peptide (5 mg/kg body weight) and the rectus abdominis muscles collected on post-burn days 1, 3, and 7. The tissues were subjected to various biochemical and immunohistochemical analyses. Treatment with SS-31 peptide prevented burn-induced increases in the caspase 3 activity (p < 0.05) and apoptosis (p < 0.01) on post-burn day 7. The SS-31 peptide treatment also prevented the increase in the expression levels of phosphatase and tensin homolog (PTEN) on post-burn days 3 and 7. Burn injury-induced increases in the levels of two ER stress markers, binding immunoglobulin protein (BiP) and protein disulfide isomerase (PDI), were significantly decreased by the SS-31 peptide treatments on post-burn day 7 and on day 3 for BiP as well (p < 0.05). The effects of SS-31 appear to be, in part, due to its ability to reduce oxidative stress in burned mice, evidenced by reduced expression of oxidized proteins that were clearly evident on post-burn day 7. Our results demonstrate a possible therapeutic potential of SS-31 peptide to ameliorate the adverse effects of burn injury in skeletal muscle. PMID:21937949

  16. Respective Contribution of Mitochondrial Superoxide and pH to Mitochondria-targeted Circularly Permuted Yellow Fluorescent Protein (mt-cpYFP) Flash Activity*

    PubMed Central

    Wei-LaPierre, Lan; Gong, Guohua; Gerstner, Brent J.; Ducreux, Sylvie; Yule, David I.; Pouvreau, Sandrine; Wang, Xianhua; Sheu, Shey-Shing; Cheng, Heping; Dirksen, Robert T.; Wang, Wang

    2013-01-01

    Superoxide flashes are transient bursts of superoxide production within the mitochondrial matrix that are detected using the superoxide-sensitive biosensor, mitochondria-targeted circularly permuted YFP (mt-cpYFP). However, due to the pH sensitivity of mt-cpYFP, flashes were suggested to reflect transient events of mitochondrial alkalinization. Here, we simultaneously monitored flashes with mt-cpYFP and mitochondrial pH with carboxy-SNARF-1. In intact cardiac myocytes and purified skeletal muscle mitochondria, robust mt-cpYFP flashes were accompanied by only a modest increase in SNARF-1 ratio (corresponding to a pH increase of <0.1), indicating that matrix alkalinization is minimal during an mt-cpYFP flash. Individual flashes were also accompanied by stepwise increases of MitoSOX signal and decreases of NADH autofluorescence, supporting the superoxide origin of mt-cpYFP flashes. Transient matrix alkalinization induced by NH4Cl only minimally influenced flash frequency and failed to alter flash amplitude. However, matrix acidification modulated superoxide flash frequency in a bimodal manner. Low concentrations of nigericin (< 100 nm) that resulted in a mild dissipation of the mitochondrial pH gradient increased flash frequency, whereas a maximal concentration of nigericin (5 μm) collapsed the pH gradient and abolished flash activity. These results indicate that mt-cpYFP flash events reflect a burst in electron transport chain-dependent superoxide production that is coincident with a modest increase in matrix pH. Furthermore, flash activity depends strongly on a combination of mitochondrial oxidation and pH gradient. PMID:23457298

  17. Differential protection by nitroxides and hydroxylamines to radiation-induced and metal ion-catalyzed oxidative damage.

    PubMed

    Xavier, Sandhya; Yamada, Ken-ichi; Samuni, Ayelet M; Samuni, Amram; DeGraff, William; Krishna, Murali C; Mitchell, James B

    2002-11-14

    Modulation of radiation- and metal ion-catalyzed oxidative-induced damage using plasmid DNA, genomic DNA, and cell survival, by three nitroxides and their corresponding hydroxylamines, were examined. The antioxidant property of each compound was independently determined by reacting supercoiled DNA with copper II/1,10-phenanthroline complex fueled by the products of hypoxanthine/xanthine oxidase (HX/XO) and noting the protective effect as assessed by agarose gel electrophoresis. The nitroxides and their corresponding hydroxylamines protected approximately to the same degree (33-47% relaxed form) when compared to 76.7% relaxed form in the absence of protectors. Likewise, protection by both the nitroxide and corresponding hydroxylamine were observed for Chinese hamster V79 cells exposed to hydrogen peroxide. In contrast, when plasmid DNA damage was induced by ionizing radiation (100 Gy), only nitroxides (10 mM) provide protection (32.4-38.5% relaxed form) when compared to radiation alone or in the presence of hydroxylamines (10 mM) (79.8% relaxed form). Nitroxide protection was concentration dependent. Radiation cell survival studies and DNA double-strand break (DBS) assessment (pulse field electrophoresis) showed that only the nitroxide protected or prevented damage, respectively. Collectively, the results show that nitroxides and hydroxylamines protect equally against the damage mediated by oxidants generated by the metal ion-catalyzed Haber-Weiss reaction, but only nitroxides protect against radiation damage, suggesting that nitroxides may more readily react with intermediate radical species produced by radiation than hydroxylamines.

  18. Ionizable Nitroxides for Studying Local Electrostatic Properties of Lipid Bilayers and Protein Systems by EPR

    PubMed Central

    Voinov, Maxim A.; Smirnov, Alex I.

    2016-01-01

    Electrostatic interactions are known to play one of the major roles in the myriad of biochemical and biophysical processes. In this Chapter we describe biophysical methods to probe local electrostatic potentials of proteins and lipid bilayer systems that is based on an observation of reversible protonation of nitroxides by EPR. Two types of the electrostatic probes are discussed. The first one includes methanethiosulfonate derivatives of protonatable nitroxides that could be used for highly specific covalent modification of the cysteine’s sulfhydryl groups. Such spin labels are very similar in magnetic parameters and chemical properties to conventional MTSL making them suitable for studying local electrostatic properties of protein-lipid interfaces. The second type of EPR probes is designed as spin-labeled phospholipids having a protonatable nitroxide tethered to the polar head group. The probes of both types report on their ionization state through changes in magnetic parameters and a degree of rotational averaging, thus, allowing one to determine the electrostatic contribution to the interfacial pKa of the nitroxide, and, therefore, determining the local electrostatic potential. Due to their small molecular volume these probes cause a minimal perturbation to the protein or lipid system while covalent attachment secure the position of the reporter nitroxides. Experimental procedures to characterize and calibrate these probes by EPR and also the methods to analyze the EPR spectra by least-squares simulations are also outlined. The ionizable nitroxide labels and the nitroxide-labeled phospholipids described so far cover an exceptionally wide pH range from ca. 2.5 to 7.0 pH units making them suitable to study a broad range of biophysical phenomena especially at the negatively charged lipid bilayer surfaces. The rationale for selecting proper electrostatically neutral interface for calibrating such probes and example of studying surface potential of lipid bilayer is

  19. Conformationally Constrained, Stable, Triplet Ground State (S = 1) Nitroxide Diradicals. Antiferromagnetic Chains of S = 1 Diradicals

    SciTech Connect

    Rajca, Andrzej; Takahashi, Masahiro; Pink, Maren; Spagnol, Gaelle; Rajca, Suchada

    2008-06-30

    Nitroxide diradicals, in which nitroxides are annelated to m-phenylene forming tricyclic benzobisoxazine-like structures, have been synthesized and characterized by X-ray crystallography, magnetic resonance (EPR and {sup 1}H NMR) spectroscopy, as well as magnetic studies in solution and in solid state. For the octamethyl derivative of benzobisoxazine nitroxide diradical, the conformationally constrained nitroxide moieties are coplanar with the m-phenylene, leading to large values of 2J (2J/k > 200 K in solution and 2J/k >> 300 K in the solid state). For the diradical, in which all ortho and para positions of the m-phenylene are sterically shielded, distortion of the nitroxide moieties from coplanarity is moderate, such that the singlet-triplet gaps remain large in both solution (2J/k > 200 K) and the solid state (2J/k {approx} 400-800 K), though an onset of thermal depopulation of the triplet ground state is detectable near room temperature. These diradicals have robust triplet ground states with strong ferromagnetic coupling and good stability at ambient conditions. Magnetic behavior of the nitroxide diradicals at low temperature is best fit to the model of one-dimensional S = 1 Heisenberg chains with intrachain antiferromagnetic coupling. The antiferromagnetic coupling between the S = 1 diradicals may be associated with the methyl nitroxide C-H {hor_ellipsis} O contacts, including nonclassical hydrogen bonds. These unprecedented organic S = 1 antiferromagnetic chains are highly isotropic, compared to those of the extensively studied Ni(II)-based chains.

  20. Antioxidant mechanism of mitochondria-targeted plastoquinone SkQ1 is suppressed in aglycemic HepG2 cells dependent on oxidative phosphorylation.

    PubMed

    Ježek, Jan; Engstová, Hana; Ježek, Petr

    2017-09-01

    Previously suggested antioxidant mechanisms for mitochondria-targeted plastoquinone SkQ1 included: i) ion-pairing of cationic SkQ1(+) with free fatty acid anions resulting in uncoupling; ii) SkQ1H2 ability to interact with lipoperoxyl radical; iii) interference with electron flow at the inner ubiquinone (Q) binding site of Complex III (Qi), involving the reduction of SkQ1 to SkQ1H2 by ubiquinol. We elucidated SkQ1 antioxidant properties by confocal fluorescence semi-quantification of mitochondrial superoxide (Jm) and cytosolic H2O2 (Jc) release rates in HepG2 cells. Only in glycolytic cells, SkQ1 prevented the rotenone-induced enhancement of Jm and Jc but not basal releases without rotenone. The effect ceased in glutaminolytic aglycemic cells, in which the redox parameter NAD(P)H/FAD increased after rotenone in contrast to its decrease in glycolytic cells. Autofluorescence decay indicated decreased NADPH/NADH ratios with rotenone in both metabolic modes. SkQ1 did not increase cell respiration and diminished Jm established high by antimycin or myxothiazol but not by stigmatellin. The revealed SkQ1 antioxidant modes reflect its reduction to SkQ1H2 at Complex I IQ or Complex III Qi site. Both reductions diminish electron diversions to oxygen thus attenuating superoxide formation. Resulting SkQ1H2 oxidizes back to SkQ1at the second (flavin) Complex I site, previously indicated for MitoQ10. Regeneration proceeds only at lower NAD(P)H/FAD in glycolytic cells. In contrast, cyclic SkQ1 reduction/SkQ1H2 oxidation does not substantiate antioxidant activity in intact cells in the absence of oxidative stress (neither pro-oxidant activity, representing a great advantage). A targeted delivery to oxidative-stressed tissues is suggested for the effective antioxidant therapy based on SkQ1. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Mitochondria-targeted esculetin inhibits PAI-1 levels by modulating STAT3 activation and miR-19b via SIRT3: Role in acute coronary artery syndrome.

    PubMed

    Katta, Sujana; Karnewar, Santosh; Panuganti, Devayani; Jerald, Mahesh Kumar; Sastry, B K S; Kotamraju, Srigiridhar

    2018-01-01

    In this study we explored the microRNAs responsible for the regulation of PAI-1 during LPS-stimulated inflammation in human aortic endothelial cells and subsequently studied the effect of a newly synthesized mitochondria-targeted esculetin (Mito-Esc) that was shown for its anti-atherosclerotic potential, in modulating PAI-1 levels and its targeted miRs during angiotensin-II-induced atherosclerosis in ApoE(-/-) mice. LPS-stimulated PAI-1 was accompanied with an upregulation of miR-19b and down-regulation of miR-30c. These effects of LPS on PAI-1 were reversed in the presence of both parent esculetin and Mito-Esc. However, the effect of Mito-Esc was more pronounced in the regulation of PAI-1. In addition, LPS-stimulated PAI-1 expression was significantly decreased in cells treated with Anti-miR-19b, thereby suggesting that miR-19b co-expression plays a key role in PAI-1 regulation. The results also show that incubation of cells with Stattic, an inhibitor of STAT-3, inhibited LPS-stimulated PAI-1 expression. Interestingly, knockdown of SIRT3, a mitochondrial biogenetic marker, enhanced PAI-1 levels via modulation of miR-19b and -30c. Mito-Esc treatment significantly inhibited Ang-II-induced PAI-1, possibly via altering miR-19b and 30c in ApoE(-/-) mice. The association between PAI-1, miR-19b and -30c were further confirmed in plasma and microparticles isolated from patients suffering from acute coronary syndrome of various degrees. Taken together, LPS-induced PAI-1 involves co-expression of miR-19b and down regulation of miR-30c, and Mito-Esc treatment by modulating miR-19b and miR-30c through SIRT3 activation, inhibits PAI-1 levels that, in part, contribute to its anti-atherosclerotic effects. Moreover, there exists a strong positive correlation between miR-19b and PAI-1 in patients suffering from ST-elevated myocardial infarction. © 2017 Wiley Periodicals, Inc.

  2. Mitochondria-targeted plastoquinone derivatives as tools to interrupt execution of the aging program. 1. Cationic plastoquinone derivatives: synthesis and in vitro studies.

    PubMed

    Antonenko, Y N; Avetisyan, A V; Bakeeva, L E; Chernyak, B V; Chertkov, V A; Domnina, L V; Ivanova, O Yu; Izyumov, D S; Khailova, L S; Klishin, S S; Korshunova, G A; Lyamzaev, K G; Muntyan, M S; Nepryakhina, O K; Pashkovskaya, A A; Pletjushkina, O Yu; Pustovidko, A V; Roginsky, V A; Rokitskaya, T I; Ruuge, E K; Saprunova, V B; Severina, I I; Simonyan, R A; Skulachev, I V; Skulachev, M V; Sumbatyan, N V; Sviryaeva, I V; Tashlitsky, V N; Vassiliev, J M; Vyssokikh, M Yu; Yaguzhinsky, L S; Zamyatnin, A A; Skulachev, V P

    2008-12-01

    concluded that SkQs are rechargeable, mitochondria-targeted antioxidants of very high efficiency and specificity. Therefore, they might be used to effectively prevent ROS-induced oxidation of lipids and proteins in the inner mitochondrial membrane in vivo.

  3. Development of nitroxide radicals-containing polymer for scavenging reactive oxygen species from cigarette smoke

    NASA Astrophysics Data System (ADS)

    Yoshitomi, Toru; Kuramochi, Kazuhiro; Binh Vong, Long; Nagasaki, Yukio

    2014-06-01

    We developed a nitroxide radicals-containing polymer (NRP), which is composed of poly(4-methylstyrene) possessing nitroxide radicals as a side chain via amine linkage, to scavenge reactive oxygen species (ROS) from cigarette smoke. In this study, the NRP was coated onto cigarette filters and its ROS-scavenging activity from streaming cigarette smoke was evaluated. The intensity of electron spin resonance signals of the NRP in the filter decreased after exposure to cigarette smoke, indicating consumption of nitroxide radicals. To evaluate the ROS-scavenging activity of the NRP-coated filter, the amount of peroxy radicals in an extract of cigarette smoke was measured using UV-visible spectrophotometry and 1,1-diphenyl-2-picrylhydrazyl (DPPH). The absorbance of DPPH at 517 nm decreased with exposure to cigarette smoke. When NRP-coated filters were used, the decrease in the absorbance of DPPH was prevented. In contrast, both poly[4-(cyclohexylamino)methylstyrene]- and poly(acrylic acid)-coated filters, which have no nitroxide radical, did not show any effect, indicating that the nitroxide radicals in the NRP scavenge the ROS in cigarette smoke. As a result, the extract of cigarette smoke passed through the NRP-coated filter has a lower cellular toxicity than smoke passed through poly[4-(cyclohexylamino)methylstyrene]- and poly(acrylic acid)-coated filters. Accordingly, NRP is a promising material for ROS scavenging from cigarette smoke.

  4. Prediction of nitroxide hyperfine coupling constants in solution from combined nanosecond scale simulations and quantum computations

    NASA Astrophysics Data System (ADS)

    Houriez, Céline; Ferré, Nicolas; Masella, Michel; Siri, Didier

    2008-06-01

    We present a combined theoretical approach based on analyzing molecular dynamics trajectories (at the nanosecond scale) generated by use of classical polarizable force fields and on quantum calculations to compute averaged hyperfine coupling constants. That method is used to estimate the constant of a prototypical nitroxide: the dimethylnitroxide. The molecule is embedded during the simulations in a cubic box containing about 500 water molecules and the molecular dynamics is generated using periodic conditions. Once the trajectories are achieved, the nitroxide and its first hydration shell molecules are extracted, and the coupling constants are computed by considering the latter aggregates by means of quantum computations. However, all the water molecules of the bulk are also accounted for during those computations by means of the electrostatic potential fitted method. Our results exhibit that in order to predict accurate and reliable coupling constants, one needs to describe carefully the out-of-plane motion of the nitroxide nitrogen and to sample trajectories with a time interval of 400 fs at least to generate an uncorrelated large set of nitroxide structures. Compared to Car-Parrinello molecular dynamics techniques, our approach can be used readily to compute hyperfine coupling constants of large systems, such as nitroxides of great size interacting with macromolecules such as proteins or polymers.

  5. Mechanism of brain protection by nitroxide radicals in experimental model of closed-head injury.

    PubMed

    Zhang, R; Shohami, E; Beit-Yannai, E; Bass, R; Trembovler, V; Samuni, A

    1998-01-15

    Reactive oxygen-derived species were previously implicated in mediation of post-traumatic brain damage; however, the efficacy of traditional antioxidants in preventing/reversing the damage is sometimes limited. The present work focused on the mechanisms underlying the neuroprotective activity of cell permeable, nontoxic, antioxidants, namely stable nitroxide radicals in an experimental model of rat closed-head injury. Brain damage was induced by the weight-drop method and the clinical status was evaluated according to a neurological severity score at 1 h and 24 h, where the difference between these scores reflects the extent of recovery. The metal chelator deferoxamine as well as three nitroxide derivatives, differing in hydrophilicity and charge, and one hydroxylamine (a reduced nitroxide) facilitated the clinical recovery and decreased the brain edema. The nitroxides, but neither the hydroxylamine nor deferoxamine, protected the integrity of the blood-brain barrier. Superoxide dismutase also improved the clinical recovery but did not affect brain edema or the blood-brain barrier. The results suggest that by switching back and forth between themselves, the nitroxide and hydroxylamine act catalytically as self-replenishing antioxidants, and protect brain tissue by terminating radical-chain reactions, oxidizing deleterious metal ions, and by removal of intracellular superoxide.

  6. Development of nitroxide radicals-containing polymer for scavenging reactive oxygen species from cigarette smoke.

    PubMed

    Yoshitomi, Toru; Kuramochi, Kazuhiro; Binh Vong, Long; Nagasaki, Yukio

    2014-06-01

    We developed a nitroxide radicals-containing polymer (NRP), which is composed of poly(4-methylstyrene) possessing nitroxide radicals as a side chain via amine linkage, to scavenge reactive oxygen species (ROS) from cigarette smoke. In this study, the NRP was coated onto cigarette filters and its ROS-scavenging activity from streaming cigarette smoke was evaluated. The intensity of electron spin resonance signals of the NRP in the filter decreased after exposure to cigarette smoke, indicating consumption of nitroxide radicals. To evaluate the ROS-scavenging activity of the NRP-coated filter, the amount of peroxy radicals in an extract of cigarette smoke was measured using UV-visible spectrophotometry and 1,1-diphenyl-2-picrylhydrazyl (DPPH). The absorbance of DPPH at 517 nm decreased with exposure to cigarette smoke. When NRP-coated filters were used, the decrease in the absorbance of DPPH was prevented. In contrast, both poly[4-(cyclohexylamino)methylstyrene]- and poly(acrylic acid)-coated filters, which have no nitroxide radical, did not show any effect, indicating that the nitroxide radicals in the NRP scavenge the ROS in cigarette smoke. As a result, the extract of cigarette smoke passed through the NRP-coated filter has a lower cellular toxicity than smoke passed through poly[4-(cyclohexylamino)methylstyrene]- and poly(acrylic acid)-coated filters. Accordingly, NRP is a promising material for ROS scavenging from cigarette smoke.

  7. Preparation of photoactive polymers and postmodification via nitroxide trapping under UV irradiation.

    PubMed

    Mardyukov, Artur; Studer, Armido

    2013-01-11

    New types of photoactive homo and block copolymers bearing α-hydroxyalkylphenylketone (2-hydroxy-2-methyl-1-phenylpropan-1-one) moieties as backbone substituents are prepared using nitroxide-mediated radical polymerization (NMP). Such polymers can be readily activated via the Norrish-type I photoreaction to give polymeric acyl radicals. Photolysis in the presence of a persistent nitroxide, which serves as a C- radical trapping reagent, leads to chemically modified polymers conjugated with nitroxide moieties. The number-average molecular weight (M(n)) of the prepolymers and the chemically modified polymers was determined by gel permeation chromatography (GPC). Structures were further confirmed by NMR spectroscopy and by attenuated total reflection (ATR) Fourier transform infrared (FTIR) spectroscopy.

  8. In vitro synthesis of nitroxide free radicals by hog liver microsomes

    SciTech Connect

    Valvis, I.I.; Lischick, D.; Shen, D.; Sofer, S.S. )

    1990-01-01

    The in vitro biooxidation of 4-hydroxy-2,2,6,6-tetra methylpiperidine (TEMP), 4-hydroxy-2,2,4,4-tetra methyl-1,3-oxazolidine (TEMO) and diphenylamine (DPA) by hog liver microsomes to their respective nitroxide free radicals, 4-hydroxy-2,2,6,6-tetra methylpiperidine-1-oxyl (TEMPO), 2,2,4,4-tetra methyl-1,3-oxazolidine-1-oxyl (TEMOO), and diphenylnitroxide (DPNO) has been investigated. For extending the life span of the liver microsomes, a calcium alginate immobilization procedure was used. The biooxidation rates of the above amines to their respective nitroxide metabolites were measured by means of oxygen uptake at 37 degrees C and pH 7.4. N-octylamine was found to be an activator in the biooxidation of the amines. The formation of the nitroxide radicals was identified by E.S.R. spectroscopy.

  9. Measuring nanometer distances in nucleic acids using a sequence-independent nitroxide probe

    PubMed Central

    Qin, Peter Z; Haworth, Ian S; Cai, Qi; Kusnetzow, Ana K; Grant, Gian Paola G; Price, Eric A; Sowa, Glenna Z; Popova, Anna; Herreros, Bruno; He, Honghang

    2008-01-01

    This protocol describes the procedures for measuring nanometer distances in nucleic acids using a nitroxide probe that can be attached to any nucleotide within a given sequence. Two nitroxides are attached to phosphorothioates that are chemically substituted at specific sites of DNA or RNA. Inter-nitroxide distances are measured using a four-pulse double electron–electron resonance technique, and the measured distances are correlated to the parent structures using a Web-accessible computer program. Four to five days are needed for sample labeling, purification and distance measurement. The procedures described herein provide a method for probing global structures and studying conformational changes of nucleic acids and protein/nucleic acid complexes. PMID:17947978

  10. ROS-Responsive Mitochondria-Targeting Blended Nanoparticles: Chemo- and Photodynamic Synergistic Therapy for Lung Cancer with On-Demand Drug Release upon Irradiation with a Single Light Source.

    PubMed

    Yue, Caixia; Yang, Yuming; Zhang, Chunlei; Alfranca, Gabriel; Cheng, Shangli; Ma, Lijun; Liu, Yanlei; Zhi, Xiao; Ni, Jian; Jiang, Weihua; Song, Jie; de la Fuente, Jesús M; Cui, Daxiang

    2016-01-01

    Mitochondria in cancer cells maintain a more negative membrane potential than normal cells. Mitochondria are the primary source of cellular reactive oxygen species (ROS), which are necessary for photodynamic therapy. Thus, the strategy of targeting mitochondria can maximize the photodynamic therapeutic efficiency for cancer. Here we report, for the first time, synthesis of a new mitochondria-targeting drug delivery system, ZnPc/CPT-TPPNPs. To synthesize this novel compound, polyethylene glycol was functionalized with thioketal linker-modified camptothecin (TL-CPT) and triphenylphosphonium to form the block copolymer, TL-CPT-PEG1K-TPP. The ZnPc/CPT-TPPNPs was constructed for delivery of the photosensitizer Zinc phthalocyanine (ZnPc) by blending the block copolymer TL-CPT-PEG1K-TPP with 1, 2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethylene glycol)] (DSPE-PEG).Triphenylphosphine can accumulate selectively several hundred-fold within mitochondria. The thioketal linker is ROS-responsive and CPT can be released upon ROS cleavage. We also show that the ZnPc loaded in ZnPc/CPT-TPPNPs absorbed the 633 nm laser to produce ROS, which could be utilized both in photodynamic therapy and to cleave the thioketal linker thereby releasing camptothecin for chemotherapy. Thus, the mitochondria-targeting nanoparticles could elevate photodynamic therapeutic efficacy. Our results showed that surface modification of the nanoparticles with triphenylphosphine cations facilitated efficient subcellular delivery of the photosensitizer to mitochondria. The nanoparticles had a good ROS-responsive effect to release CPT, which could transfer to the nucleus and interfere with DNA replication as a topoisomeraseⅠinhibitor. Thus, the blended nanoparticles provide a new promising approach as a mitochondria-targeting ROS-activated chemo- and photodynamic therapy with a single light source for lung cancer.

  11. ROS-Responsive Mitochondria-Targeting Blended Nanoparticles: Chemo- and Photodynamic Synergistic Therapy for Lung Cancer with On-Demand Drug Release upon Irradiation with a Single Light Source

    PubMed Central

    Yue, Caixia; Yang, Yuming; Zhang, Chunlei; Alfranca, Gabriel; Cheng, Shangli; Ma, Lijun; Liu, Yanlei; Zhi, Xiao; Ni, Jian; Jiang, Weihua; Song, Jie; de la Fuente, Jesús M.; Cui, Daxiang

    2016-01-01

    Mitochondria in cancer cells maintain a more negative membrane potential than normal cells. Mitochondria are the primary source of cellular reactive oxygen species (ROS), which are necessary for photodynamic therapy. Thus, the strategy of targeting mitochondria can maximize the photodynamic therapeutic efficiency for cancer. Here we report, for the first time, synthesis of a new mitochondria-targeting drug delivery system, ZnPc/CPT-TPPNPs. To synthesize this novel compound, polyethylene glycol was functionalized with thioketal linker-modified camptothecin (TL-CPT) and triphenylphosphonium to form the block copolymer, TL-CPT-PEG1K-TPP. The ZnPc/CPT-TPPNPs was constructed for delivery of the photosensitizer Zinc phthalocyanine (ZnPc) by blending the block copolymer TL-CPT-PEG1K-TPP with 1, 2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethylene glycol)] (DSPE-PEG).Triphenylphosphine can accumulate selectively several hundred-fold within mitochondria. The thioketal linker is ROS-responsive and CPT can be released upon ROS cleavage. We also show that the ZnPc loaded in ZnPc/CPT-TPPNPs absorbed the 633 nm laser to produce ROS, which could be utilized both in photodynamic therapy and to cleave the thioketal linker thereby releasing camptothecin for chemotherapy. Thus, the mitochondria-targeting nanoparticles could elevate photodynamic therapeutic efficacy. Our results showed that surface modification of the nanoparticles with triphenylphosphine cations facilitated efficient subcellular delivery of the photosensitizer to mitochondria. The nanoparticles had a good ROS-responsive effect to release CPT, which could transfer to the nucleus and interfere with DNA replication as a topoisomeraseⅠinhibitor. Thus, the blended nanoparticles provide a new promising approach as a mitochondria-targeting ROS-activated chemo- and photodynamic therapy with a single light source for lung cancer. PMID:27877240

  12. Stability of SG1 nitroxide towards unprotected sugar and lithium salts: a preamble to cellulose modification by nitroxide-mediated graft polymerization.

    PubMed

    Moreira, Guillaume; Charles, Laurence; Major, Mohamed; Vacandio, Florence; Guillaneuf, Yohann; Lefay, Catherine; Gigmes, Didier

    2013-01-01

    The range of applications of cellulose, a glucose-based polysaccharide, is limited by its inherently poor mechanical properties. The grafting of synthetic polymer chains by, for example, a "grafting from" process may provide the means to broaden the range of applications. The nitroxide-mediated polymerization (NMP) method is a technique of choice to control the length, the composition and the architecture of the grafted copolymers. Nevertheless, cellulose is difficult to solubilize in organic media because of inter- and intramolecular hydrogen bonds. One possibility to circumvent this limitation is to solubilize cellulose in N,N-dimethylformamide (DMF) or N,N-dimethylacetamide (DMA) with 5 to 10 wt % of lithium salts (LiCl or LiBr), and carry out grafted polymerization in this medium. The stability of nitroxides such as SG1 has not been studied under these conditions yet, even though these parameters are of crucial importance to perform the graft modification of polysaccharide by NMP. The aim of this work is to offer a model study of the stability of the SG1 nitroxide in organic media in the presence of unprotected glucose or cellobiose (used as a model of cellulose) and in the presence of lithium salts (LiBr or LiCl) in DMF or DMA. Contrary to TEMPO, SG1 proved to be stable in the presence of unprotected sugar, even with an excess of 100 molar equivalents of glucose. On the other hand, lithium salts in DMF or DMA clearly degrade SG1 nitroxide as proven by electron-spin resonance measurements. The instability of SG1 in these lithium-containing solvents may be explained by the acidification of the medium by the hydrolysis of DMA in the presence of LiCl. This, in turn, enables the disproportionation of the SG1 nitroxide into an unstable hydroxylamine and an oxoammonium ion. Once the conditions to perform an SG1-based nitroxide-mediated graft polymerization from cellobiose have been established, the next stage of this work will be the modification of cellulose and

  13. Anti-inflammatory, analgesic and antioxidant activities of novel kyotorphin-nitroxide hybrid molecules.

    PubMed

    Bi, Wei; Bi, Yue; Gao, Xiang; Yan, Xin; Zhang, Yanrong; Xue, Ping; Bammert, Catherine E; Legalley, Thomas D; Michael Gibson, K; Bi, Lanrong; Wang, Jia-Xiang

    2016-04-15

    Mitochondrial oxidative damage contributes to a wide range of pathologies, including ischemia/reperfusion (I/R) injury, cardiovascular disorders and neurodegenerative diseases. Accordingly, protecting mitochondria from oxidative damage should possess therapeutic relevance. In the present study, we have designed and synthesized a series of novel kyotorphin-nitroxide hybrid molecules, and examined their free radical scavenging activities, in addition to their anti-inflammatory and analgesic activities. We have further characterized these compounds in a simulated I/R cellular model. Our findings suggest that the protective effects of kyotorphin-nitroxides partially reside in maintaining optimal mitochondrial function. Published by Elsevier Ltd.

  14. First-principles study of polyacetylene derivatives bearing nitroxide radicals

    NASA Astrophysics Data System (ADS)

    Bilgiç, Beyza; Kılıç, Çetin; Esat, Burak

    2011-09-01

    Electrodes made of organic polymers bearing redox-active radical pendant groups have attractive features for use in rechargeable batteries. Electronic structure and electrochemical properties of cathode- and anode-active organic polymers are investigated here by means of first-principles calculations performed in the framework of the density functional theory. We consider organic radical polymers (ORPs) that consist of trans-polyacetylene derivatives bearing a variety of nitroxide radicals. A number of neutral and charged supercells are utilized to compute the ionization potentials and electron affinities as well as the one-electron states of these ORPs. By revealing the polyacetylene-derived highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) as well as the radical-derived singly occupied molecular orbital (SOMO), the variation of the SOMO energy within the HOMO-LUMO gap is determined in the course of the oxidization or reduction of ORPs. Our results indicate that the ionization potential I and electron affinity A of polyacetylene would act as a lower or upper bound in the variation of the electrochemical potential of cathode- or anode-active ORPs in the course of battery discharge or charge owing to pinning of the radical-derived SOMO to the polyacetylene-derived HOMO or LUMO. Accordingly, it is anticipated that the electrochemical “window” [-I,-A] of the polymeric backbone of ORPs will impose certain limitations in accomplishing a high charge/discharge voltage range in a totally organic rechargeable battery with positive and negative electrodes made of cathode- and anode-active ORPs, respectively. On the other hand, our findings suggest that one could, in principle, take advantage of using two different (conducting) polymeric backbones in the anode and cathode with adjusted HOMO and LUMO offsets once the electron transfer is accomplished to take place through the conducting backbones.

  15. Site-specific immobilization of proteins at zeolite L crystals by nitroxide exchange reactions.

    PubMed

    Becker, Maike; De Cola, Luisa; Studer, Armido

    2011-03-28

    Site-selective immobilization of dyes and different protein recognizing entities at the surface of zeolite L crystals using mild radical nitroxide exchange reactions is reported. Exposure of these crystals to aqueous protein solutions leads to site-selective immobilization of proteins onto the crystals.

  16. Spin-state-correlated optical properties of copper(ii)-nitroxide based molecular magnets.

    PubMed

    Barskaya, Irina Yu; Veber, Sergey L; Suturina, Elizaveta A; Sherin, Peter S; Maryunina, Kseniya Yu; Artiukhova, Natalia A; Tretyakov, Evgeny V; Sagdeev, Renad Z; Ovcharenko, Victor I; Gritsan, Nina P; Fedin, Matvey V

    2017-10-03

    Molecular magnets based on copper(ii) ions and stable nitroxide radicals exhibit promising switchable behavior triggered by a number of external stimuli; however, their spin-state-correlated optical properties vital for photoinduced switching have not been profoundly investigated to date. Herein, the electronic absorption spectra of single crystals of three representatives of this unique family are studied experimentally and theoretically in the visible and near-IR regions. We established that the color of the complexes is mainly determined by optical properties of the nitroxide radicals, whereas the Cu(hfac)2 fragment contributes to the near-IR range with the intensity smaller by an order of magnitude. The thermochromism of these complexes evident upon thermal spin state switching is mainly caused by a spectral shift of the absorption bands of the nitroxides. The vibrational progression observed in the visible range for single crystals as well as for solutions of pure nitroxides is well reproduced by DFT calculations, where the C-C stretching mode governs the observed progression. The analysis of the spectra of single crystals in the near-IR region reveals changes in the energy and in the intensity of the copper(ii) d-d transitions, which are well reproduced by SOC-NEVPT2 calculations and owe to the flip of the Jahn-Teller axis in the coordination environment of copper. Further strategies for designing bidirectional magnetic photoswitches using these appealing compounds are discussed.

  17. A high saturation factor in Overhauser DNP with nitroxide derivatives: the role of (14)N nuclear spin relaxation.

    PubMed

    Enkin, Nikolay; Liu, Guoquan; Gimenez-Lopez, Maria Del Carmen; Porfyrakis, Kyriakos; Tkach, Igor; Bennati, Marina

    2015-05-07

    Overhauser DNP enhancements of toluene were measured at a magnetic field of 0.35 Tesla in a series of chemically functionalized nitroxide radicals. We observe that the enhancements increase systematically with polarizer size and rotational correlation time. Examination of the saturation factor of (14)N nitroxides by pulsed ELDOR spectroscopy led to a quantitative interpretation of the enhancements, for which the saturation factor increases up to almost unity due to enhanced nuclear ((14)N) relaxation in the nitroxide radical. The observation has a direct impact on the choice of optimum DNP polarizers in liquids.

  18. A new model for Overhauser enhanced nuclear magnetic resonance using nitroxide radicals.

    PubMed

    Armstrong, Brandon D; Han, Songi

    2007-09-14

    Nitroxide free radicals are the most commonly used source for dynamic nuclear polarization (DNP) enhanced nuclear magnetic resonance (NMR) experiments and are also exclusively employed as spin labels for electron spin resonance (ESR) spectroscopy of diamagnetic molecules and materials. Nitroxide free radicals have been shown to have strong dipolar coupling to (1)H in water, and thus result in large DNP enhancement of (1)H NMR signal via the well known Overhauser effect. The fundamental parameter in a DNP experiment is the coupling factor, since it ultimately determines the maximum NMR signal enhancements which can be achieved. Despite their widespread use, measurements of the coupling factor of nitroxide free radicals have been inconsistent, and current models have failed to successfully explain our experimental data. We found that the inconsistency in determining the coupling factor arises from not taking into account the characteristics of the ESR transitions, which are split into three (or two) lines due to the hyperfine coupling of the electron to the (14)N nuclei (or (15)N) of the nitric oxide radical. Both intermolecular Heisenberg spin exchange interactions as well as intramolecular nitrogen nuclear spin relaxation mix the three (or two) ESR transitions. However, neither effect has been taken into account in any experimental studies on utilizing or quantifying the Overhauser driven DNP effects. The expected effect of Heisenberg spin exchange on Overhauser enhancements has already been theoretically predicted and observed by Bates and Drozdoski [J. Chem. Phys. 67, 4038 (1977)]. Here, we present a new model for quantifying Overhauser enhancements through nitroxide free radicals that includes both effects on mixing the ESR hyperfine states. This model predicts the maximum saturation factor to be considerably higher by the effect of nitrogen nuclear spin relaxation. Because intramolecular nitrogen spin relaxation is independent of the nitroxide concentration

  19. Effectiveness of Analogs of the GS-Nitroxide, JP4-039, as Total Body Irradiation Mitigators

    PubMed Central

    W. EPPERLY, MICHAEL; R. SACHER, JOSHUA; KRAINZ, TANJA; ZHANG, XIAOLIN; WIPF, PETER; LIANG, MARY; FISHER, RENEE; LI, SONG; WANG, HONG; S. GREENBERGER, JOEL

    2017-01-01

    Background/Aim: Mitochondrial-targeted gramicidin S (GS)-nitroxide, JP4-039, has been demonstrated to be a potent radiation mitigator, and safe over a wide dose range. In addition, JP4-039 has organ-specific effectiveness when locally applied. Materials and Methods: We tested the effect of another GS-nitroxide, XJB-5-131, which has more effective mitochondrial localization, and compared these results to those for radiation mitigation against the hematopoietic syndrome, and two analogs of JP4-039, which have the same mitochondrial localization signal, but different chemical payloads: JRS527.084 contains a second nitroxide per molecule, and TK649.030 contains an ester group attached to the nitroxide. Results: The results demonstrate the superiority of JP4-039 as a systemic radiation mitigator. Conclusion: Structure–activity relationships and bioassays demonstrate that JP4-039 is an optimized small-molecule radiation mitigator. PMID:28064218

  20. Long-term administration of the mitochondria-targeted antioxidant mitoquinone mesylate fails to attenuate age-related oxidative damage or rescue the loss of muscle mass and function associated with aging of skeletal muscle.

    PubMed

    Sakellariou, Giorgos K; Pearson, Timothy; Lightfoot, Adam P; Nye, Gareth A; Wells, Nicola; Giakoumaki, Ifigeneia I; Griffiths, Richard D; McArdle, Anne; Jackson, Malcolm J

    2016-11-01

    Age-related skeletal muscle dysfunction is the underlying cause of morbidity that affects up to half the population aged 80 and over. Considerable evidence indicates that oxidative damage and mitochondrial dysfunction contribute to the sarcopenic phenotype that occurs with aging. To examine this, we administered the mitochondria-targeted antioxidant mitoquinone mesylate {[10-(4,5-dimethoxy-2-methyl-3,6-dioxo-1,4-cyclohexadien-1-yl)decyl] triphenylphosphonium; 100 μM} to wild-type C57BL/6 mice for 15 wk (from 24 to 28 mo of age) and investigated the effects on age-related loss of muscle mass and function, changes in redox homeostasis, and mitochondrial organelle integrity and function. We found that mitoquinone mesylate treatment failed to prevent age-dependent loss of skeletal muscle mass associated with myofiber atrophy or alter a variety of in situ and ex vivo muscle function analyses, including maximum isometric tetanic force, decline in force after a tetanic fatiguing protocol, and single-fiber-specific force. We also found evidence that long-term mitoquinone mesylate administration did not reduce mitochondrial reactive oxygen species or induce significant changes in muscle redox homeostasis, as assessed by changes in 4-hydroxynonenal protein adducts, protein carbonyl content, protein nitration, and DNA damage determined by the content of 8-hydroxydeoxyguanosine. Mitochondrial membrane potential, abundance, and respiration assessed in permeabilized myofibers were not significantly altered in response to mitoquinone mesylate treatment. Collectively, these findings demonstrate that long-term mitochondria-targeted mitoquinone mesylate administration failed to attenuate age-related oxidative damage in skeletal muscle of old mice or provide any protective effect in the context of muscle aging.-Sakellariou, G. K., Pearson, T., Lightfoot, A. P., Nye, G. A., Wells, N., Giakoumaki, I. I., Griffiths, R. D., McArdle, A., Jackson, M. J. Long-term administration of the

  1. Long-term administration of the mitochondria-targeted antioxidant mitoquinone mesylate fails to attenuate age-related oxidative damage or rescue the loss of muscle mass and function associated with aging of skeletal muscle

    PubMed Central

    Sakellariou, Giorgos K.; Pearson, Timothy; Lightfoot, Adam P.; Nye, Gareth A.; Wells, Nicola; Giakoumaki, Ifigeneia I.; Griffiths, Richard D.; McArdle, Anne; Jackson, Malcolm J.

    2016-01-01

    Age-related skeletal muscle dysfunction is the underlying cause of morbidity that affects up to half the population aged 80 and over. Considerable evidence indicates that oxidative damage and mitochondrial dysfunction contribute to the sarcopenic phenotype that occurs with aging. To examine this, we administered the mitochondria-targeted antioxidant mitoquinone mesylate {[10-(4,5-dimethoxy-2-methyl-3,6-dioxo-1,4-cyclohexadien-1-yl)decyl] triphenylphosphonium; 100 μM} to wild-type C57BL/6 mice for 15 wk (from 24 to 28 mo of age) and investigated the effects on age-related loss of muscle mass and function, changes in redox homeostasis, and mitochondrial organelle integrity and function. We found that mitoquinone mesylate treatment failed to prevent age-dependent loss of skeletal muscle mass associated with myofiber atrophy or alter a variety of in situ and ex vivo muscle function analyses, including maximum isometric tetanic force, decline in force after a tetanic fatiguing protocol, and single-fiber-specific force. We also found evidence that long-term mitoquinone mesylate administration did not reduce mitochondrial reactive oxygen species or induce significant changes in muscle redox homeostasis, as assessed by changes in 4-hydroxynonenal protein adducts, protein carbonyl content, protein nitration, and DNA damage determined by the content of 8-hydroxydeoxyguanosine. Mitochondrial membrane potential, abundance, and respiration assessed in permeabilized myofibers were not significantly altered in response to mitoquinone mesylate treatment. Collectively, these findings demonstrate that long-term mitochondria-targeted mitoquinone mesylate administration failed to attenuate age-related oxidative damage in skeletal muscle of old mice or provide any protective effect in the context of muscle aging.—Sakellariou, G. K., Pearson, T., Lightfoot, A. P., Nye, G. A., Wells, N., Giakoumaki, I. I., Griffiths, R. D., McArdle, A., Jackson, M. J. Long-term administration of the

  2. In vivo EPR oximetry using an isotopically-substituted nitroxide: Potential for quantitative measurement of tissue oxygen

    PubMed Central

    Weaver, John; Burks, Scott R.; Liu, Ke Jian; Kao, Joseph P.Y.; Rosen, Gerald M.

    2017-01-01

    Variations in brain oxygen (O2) concentration can have profound effects on brain physiology. Thus, the ability to quantitate local O2 concentrations noninvasively in vivo could significantly enhance understanding of several brain pathologies. However, quantitative O2 mapping in the brain has proven difficult. The electron paramagnetic resonance (EPR) spectra of nitroxides are sensitive to molecular O2 and can be used to estimate O2 concentrations in aqueous media. We recently synthesized labile-ester-containing nitroxides, such as 3-acetoxymethoxycarbonyl-2,2,5,5-tetramethyl-1-pyrrolidinyloxyl (nitroxide 4), which accumulate in cerebral tissue after in situ hydrolysis, and thus enable spatial mapping of O2 concentrations in the mouse brain by EPR imaging. In an effort to improve O2 quantitation, we prepared 3-acetoxymethox ycarbonyl-2,2,5,5-tetra(2H3)methyl-1-(3,4,4-2H3,1-15N)pyrrolidinyloxyl (nitroxide 2), which proved to be a more sensitive probe than its normo-isotopic version for quantifying O2 in aqueous solutions of various O2 concentrations. We now demonstrate that this isotopically substituted nitroxide is ~2-fold more sensitive in vivo than the normo-isotopic nitroxide 4. Moreover, in vitro and in vivo EPR spectral-spatial imaging results with nitroxide 2 demonstrate significant improvement in resolution, reconstruction and spectral response to local O2 concentrations in cerebral tissue. Thus, isotopic-substituted nitroxides, such as 2, are excellent sensors for in vivo O2 quantitation in tissues, such as the brain. PMID:27567323

  3. Light-Induced Spin State Switching and Relaxation in Spin Pairs of Copper(II)-Nitroxide Based Molecular Magnets.

    PubMed

    Tumanov, Sergey V; Veber, Sergey L; Tolstikov, Svyatoslav E; Artiukhova, Natalia A; Romanenko, Galina V; Ovcharenko, Victor I; Fedin, Matvey V

    2017-10-02

    Similar to spin-crossover (SCO) compounds, spin states of copper(II)-nitroxide based molecular magnets can be switched by various external stimuli including temperature and light. Although photoswitching and reverse relaxation of nitroxide-copper(II)-nitroxide triads were investigated in some detail, similar study for copper(II)-nitroxide spin pairs was still missing. In this work we address photoswitching and relaxation phenomena in exchange-coupled spin pairs of this family of molecular magnets. Using electron paramagnetic resonance (EPR) spectroscopy with photoexcitation, we demonstrate that compared to triad-containing compounds the photoinduced weakly coupled spin (WS) states of copper(II)-nitroxide pairs are remarkably more stable at cryogenic temperatures and relax to the ground strongly coupled spin (SS) states on the scale of days. The structural changes between SS and WS states, e.g., differences in Cu-Onitroxide distances, are much more pronounced for spin pairs than for spin triads in most of the studied copper(II)-nitroxide based molecular magnets. This results in higher energy barrier between WS and SS states of spin pairs and governs higher stability of their photoinduced WS states. Therefore, the longer-lived photoinduced states in copper(II)-nitroxide molecular magnets should be searched within the compounds experiencing largest structural changes upon thermal spin transition. This advancement in understanding of LIESST-like phenomena in copper(II)-nitroxide molecular magnets allows us to propose them as interesting playgrounds for benchmarking the basic factors governing the stability of photoinduced states in other SCO and SCO-like photoswitchable systems.

  4. In vivo EPR oximetry using an isotopically-substituted nitroxide: Potential for quantitative measurement of tissue oxygen

    NASA Astrophysics Data System (ADS)

    Weaver, John; Burks, Scott R.; Liu, Ke Jian; Kao, Joseph P. Y.; Rosen, Gerald M.

    2016-10-01

    Variations in brain oxygen (O2) concentration can have profound effects on brain physiology. Thus, the ability to quantitate local O2 concentrations noninvasively in vivo could significantly enhance understanding of several brain pathologies. However, quantitative O2 mapping in the brain has proven difficult. The electron paramagnetic resonance (EPR) spectra of nitroxides are sensitive to molecular O2 and can be used to estimate O2 concentrations in aqueous media. We recently synthesized labile-ester-containing nitroxides, such as 3-acetoxymethoxycarbonyl-2,2,5,5-tetramethyl-1-pyrrolidinyloxyl (nitroxide 4), which accumulate in cerebral tissue after in situ hydrolysis, and thus enable spatial mapping of O2 concentrations in the mouse brain by EPR imaging. In an effort to improve O2 quantitation, we prepared 3-acetoxymethoxycarbonyl-2,2,5,5-tetra(2H3)methyl-1-(3,4,4-2H3,1-15N)pyrrolidinyloxyl (nitroxide 2), which proved to be a more sensitive probe than its normo-isotopic version for quantifying O2 in aqueous solutions of various O2 concentrations. We now demonstrate that this isotopically substituted nitroxide is ∼2-fold more sensitive in vivo than the normo-isotopic nitroxide 4. Moreover, in vitro and in vivo EPR spectral-spatial imaging results with nitroxide 2 demonstrate significant improvement in resolution, reconstruction and spectral response to local O2 concentrations in cerebral tissue. Thus, isotopic-substituted nitroxides, such as 2, are excellent sensors for in vivo O2 quantitation in tissues, such as the brain.

  5. Room-temperature electron spin relaxation of nitroxides immobilized in trehalose: Effect of substituents adjacent to NO-group.

    PubMed

    Kuzhelev, Andrey A; Strizhakov, Rodion K; Krumkacheva, Olesya A; Polienko, Yuliya F; Morozov, Denis A; Shevelev, Georgiy Yu; Pyshnyi, Dmitrii V; Kirilyuk, Igor A; Fedin, Matvey V; Bagryanskaya, Elena G

    2016-05-01

    Trehalose has been recently promoted as efficient immobilizer of biomolecules for room-temperature EPR studies, including distance measurements between attached nitroxide spin labels. Generally, the structure of nitroxide influences the electron spin relaxation times, being crucial parameters for room-temperature pulse EPR measurements. Therefore, in this work we investigated a series of nitroxides with different substituents adjacent to NO-moiety including spirocyclohexane, spirocyclopentane, tetraethyl and tetramethyl groups. Electron spin relaxation times (T1, Tm) of these radicals immobilized in trehalose were measured at room temperature at X- and Q-bands (9/34GHz). In addition, a comparison was made with the corresponding relaxation times in nitroxide-labeled DNA immobilized in trehalose. In all cases phase memory times Tm were close to 700ns and did not essentially depend on structure of substituents. Comparison of temperature dependences of Tm at T=80-300K shows that the benefit of spirocyclohexane substituents well-known at medium temperatures (∼100-180K) becomes negligible at 300K. Therefore, unless there are specific interactions between spin labels and biomolecules, the room-temperature value of Tm in trehalose is weakly dependent on the structure of substituents adjacent to NO-moiety of nitroxide. The issues of specific interactions and stability of nitroxide labels in biological media might be more important for room temperature pulsed dipolar EPR than differences in intrinsic spin relaxation of radicals.

  6. Optimization of labile esters for esterase–assisted accumulation of nitroxides into cells: A model for in vivo EPR imaging

    PubMed Central

    Burks, Scott R.; Ni, Jiahong; Muralidharan, Sukumaran; Coop, Andrew; Kao, Joseph P.Y.; Rosen, Gerald M.

    2009-01-01

    Nitroxide–based electron paramagnetic resonance (EPR) imaging agents are useful quantitative probes of O2 concentration in vivo in real time. Lipophilic, labile alkanoyloxymethyl esters of nitroxides can cross the blood-brain barrier, and after hydrolysis, the corresponding anionic nitroxide is intracellularly entrapped at levels sufficient to permit O2 measurements. The utility of nitroxides as EPR imaging agents depends critically on their ability to accumulate in the brain to high levels. In this study, we systematically investigated the relationship between the structure of the alkanoyl moiety and the ability of the corresponding labile ester to deliver nitroxide intracellularly. We demonstrate, in a cultured cell model, that for nitroxide labile esters with unbranched alkanoyl chains, increasing the chain length improves intracellular loading. Moreover, by studying an isomeric series of labile esters, we conclude that branching of the alkanoyl chain drastically reduces intracellular loading. These structural insights improve our general ability to use labile esters to deliver carboxylates intracellularly, and suggest a strategy for how to enhance delivery of nitroxide imaging agents across the blood-brain barrier in a living animal. PMID:18783260

  7. Structural Origins of Nitroxide Side Chain Dynamics on Membrane Protein [alpha]-Helical Sites

    SciTech Connect

    Kroncke, Brett M.; Horanyi, Peter S.; Columbus, Linda

    2010-12-07

    Understanding the structure and dynamics of membrane proteins in their native, hydrophobic environment is important to understanding how these proteins function. EPR spectroscopy in combination with site-directed spin labeling (SDSL) can measure dynamics and structure of membrane proteins in their native lipid environment; however, until now the dynamics measured have been qualitative due to limited knowledge of the nitroxide spin label's intramolecular motion in the hydrophobic environment. Although several studies have elucidated the structural origins of EPR line shapes of water-soluble proteins, EPR spectra of nitroxide spin-labeled proteins in detergents or lipids have characteristic differences from their water-soluble counterparts, suggesting significant differences in the underlying molecular motion of the spin label between the two environments. To elucidate these differences, membrane-exposed {alpha}-helical sites of the leucine transporter, LeuT, from Aquifex aeolicus, were investigated using X-ray crystallography, mutational analysis, nitroxide side chain derivatives, and spectral simulations in order to obtain a motional model of the nitroxide. For each crystal structure, the nitroxide ring of a disulfide-linked spin label side chain (R1) is resolved and makes contacts with hydrophobic residues on the protein surface. The spin label at site I204 on LeuT makes a nontraditional hydrogen bond with the ortho-hydrogen on its nearest neighbor F208, whereas the spin label at site F177 makes multiple van der Waals contacts with a hydrophobic pocket formed with an adjacent helix. These results coupled with the spectral effect of mutating the i {+-} 3, 4 residues suggest that the spin label has a greater affinity for its local protein environment in the low dielectric than on a water-soluble protein surface. The simulations of the EPR spectra presented here suggest the spin label oscillates about the terminal bond nearest the ring while maintaining weak contact

  8. Enzymatically Shifting Nitroxides for EPR Spectroscopy and Overhauser-Enhanced Magnetic Resonance Imaging.

    PubMed

    Audran, Gérard; Bosco, Lionel; Brémond, Paul; Franconi, Jean-Michel; Koonjoo, Neha; Marque, Sylvain R A; Massot, Philippe; Mellet, Philippe; Parzy, Elodie; Thiaudière, Eric

    2015-11-02

    In vivo investigations of enzymatic processes using non-invasive approaches are a long-lasting challenge. Recently, we showed that Overhauser-enhanced MRI is suitable to such a purpose. A β-phosphorylated nitroxide substrate prototype exhibiting keto-enol equilibrium upon enzymatic activity has been prepared. Upon enzymatic hydrolysis, a large variation of the phosphorus hyperfine coupling constant (Δa(P)=4 G) was observed. The enzymatic activities of several enzymes were conveniently monitored by electronic paramagnetic resonance (EPR). Using a 0.2 T MRI machine, in vitro and in vivo OMRI experiments were successfully performed, affording a 1200% enhanced MRI signal in vitro, and a 600% enhanced signal in vivo. These results highlight the enhanced imaging potential of these nitroxides upon specific enzymatic substrate-to-product conversion. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Oxidation of Annelated Diarylamines: Analysis of Reaction Pathways to Nitroxide Diradical and Spirocyclic Products

    SciTech Connect

    Rajca, Andrzej; Shiraishi, Kouichi; Boraty; #324; ski, Przemyslaw J.; Pink, Maren; Miyasaka, Makoto; Rajca, Suchada

    2012-02-06

    Oxidation of diaryldiamine 2, a tetrahydrodiazapentacene derivative, provides diarylnitroxide diradical 1 accompanied by an intermediate nitroxide monoradical and a multitude of isolable diamagnetic products. DFT-computed tensors for EPR spectra and paramagnetic {sup 1}H NMR isotropic shifts for nitroxide diradical 1 show good agreement with the experimental EPR spectra in rigid matrices and paramagnetic {sup 1}H NMR spectra in solution, respectively. Examination of the diamagnetic products elucidates their formation via distinct pathways involving C-O bond-forming reactions, including Baeyer-Villiger-type oxidations. An unusual diiminoketone structure and two spirocyclic structures of the predominant diamagnetic products are confirmed by either X-ray crystallography or correlations between DFT-computed and experimental spectroscopic data such as {sup 1}H, {sup 13}C, and {sup 15}N NMR chemical shifts and electronic absorption spectra.

  10. Nitroxide polymer networks formed by Michael addition: on site-cured electrode-active organic coating.

    PubMed

    Ibe, Takeshi; Frings, Rainer B; Lachowicz, Artur; Kyo, Soichi; Nishide, Hiroyuki

    2010-05-28

    Highly and homogeneously crosslinked poly(beta-ketoester) networks densely bearing robust nitroxide radicals were prepared via a click-type and stepwise Michael polyaddition. A half-battery cell composed of the thermally-cured radical network coatings displayed a rapid, reversible, and almost stoichiometric redox-activity even with a thickness of ca. 10 mum, which may be applicable as the electrode of organic-based rechargeable devices.

  11. Rare Earth Metal Complexes of Bidentate Nitroxide Ligands: Synthesis and Electrochemistry.

    PubMed

    Kim, Jee Eon; Bogart, Justin A; Carroll, Patrick J; Schelter, Eric J

    2016-01-19

    We report rare earth metal complexes with tri- and bidentate ligands including strongly electron-donating nitroxide groups. The tridentate ligand 1,3,5-tris(2'-tert-butylhydroxylaminoaryl)benzene (H3arene-triNOx) was complexed to cerium(IV) in a 2:1 ligand-to-metal stoichiometry as Ce(Harene-triNOx)2 (1). Cyclic voltammetry of this compound showed stabilization of the tetravalent cerium cation with a Ce(IV/III) couple at E1/2 = -1.82 V versus Fc/Fc(+). On the basis of the uninvolvement of the third nitroxide group in the coordination chemistry with the cerium(IV) cation, the ligand system was redesigned toward a simpler bidentate mode, and a series of rare earth metal-arene-diNOx complexes were prepared with La(III), Ce(IV), Pr(III), Tb(III), and Y(III), [RE(arene-diNOx)2](-) ([2-RE](-), RE = La, Pr, Y, Tb) and Ce(IV)(arene-diNOx)2, where H2arene-diNOx = 1,3-bis(2'-tert-butylhydroxylaminoaryl)benzene. The core structures were isostructural throughout the series, with three nitroxide groups in η(2) binding modes and one κ(1) nitroxide group coordinated to the metal center in the solid state. In all cases except Ce(IV)(arene-diNOx)2, electrochemical analysis described two subsequent, ligand-based, quasi-reversible redox waves, indicating that a stable [N-O•] group was generated on the electrochemical time scale. Chemical oxidation of the terbium complex was performed, and isolation of the resulting complex, Tb(arene-diNOx)2·CH2Cl2 (3·CH2Cl2), confirmed the assignment of the cyclic voltammograms. Magnetic data showed no evidence of mixing between the Tb(III) states and the states of the open-shell ligand.

  12. Anaerobic nitroxide-catalyzed oxidation of alcohols using the NO+/NO· redox pair.

    PubMed

    Holan, Martin; Jahn, Ullrich

    2014-01-03

    A new method for alcohol oxidation using TEMPO or AZADO in conjunction with BF3·OEt2 or LiBF4 as precatalysts and tert-butyl nitrite as a stoichiometric oxidant has been developed. The system is based on a NO(+)/NO· pair for nitroxide reoxidation under anaerobic conditions. This allows the simple, high-yielding conversion of various achiral and chiral alcohols to carbonyl compounds without epimerization and no formation of nonvolatile byproducts.

  13. Tissue redox activity as a sensing platform for imaging of cancer based on nitroxide redox cycle.

    PubMed

    Zhelev, Zhivko; Aoki, Ichio; Gadjeva, Veselina; Nikolova, Biliana; Bakalova, Rumiana; Saga, Tsuneo

    2013-04-01

    The experience in free radical biology and medicine shows the crucial role of redox signalling in carcinogenesis. The cells and tissues of healthy mammals are characterised by a low level of reactive oxygen species (ROS) and some constant (reference) level of reducing equivalents. Increasing of ROS above the critical level provokes genomic instability. The present study describes universal methodology for direct imaging of tissue redox activity in carcinogenesis, which allows a differentiation of cancer development from normal condition. The experiments were conducted on: neuroblastoma-bearing mice; colon cancer-bearing mice; and healthy mice. The tissue redox activity was visualised in vivo by nitroxide-enhanced magnetic resonance imaging (MRI) on anesthetised animals. The method is based on nitroxide redox cycle, coupled with appearance/disappearance of MRI signal. The half-life (τ1/2) of nitroxide-enhanced MRI signal in the respective tissue was used as a diagnostic marker. The study provides direct evidence that healthy and cancer-bearing mammalian tissues are characterised by different redox activities - a basis for cancer diagnosis. The tissues (cancer and 'normal') of cancer-bearing mammals were characterised by a long-lived MRI signal (τ1/2>14 min), indicating a high oxidative activity. The tissues of healthy organism were characterised by a short-lived MRI signal (τ1/2=1-3 min), indicating a high reducing activity. The study shows that tissue redox activity is a sensing platform for imaging of cancer using nitroxide-enhanced MRI. It also suggests that 'normal' tissues of cancer-bearing organism are susceptible to oxidative damage. Copyright © 2012 Elsevier Ltd. All rights reserved.

  14. Syntheses, crystal structures, magnetic and luminescence properties of five novel lanthanide complexes of nitronyl nitroxide radical

    SciTech Connect

    Wang, Ya-Li; Gao, Yuan-Yuan; Ma, Yue; Wang, Qing-Lun; Li, Li-Cun; Liao, Dai-Zheng

    2013-06-01

    Five novel Ln(III) complexes based on a new nitronyl nitroxide radical have been synthesized, characterized structurally and magnetically: [Ln(hfac)₃(NITPh-3-Br-4-OMe)₂] (Ln(III)=Eu(1), Gd(2), Tb(3), Dy(4), Ho(5); hfac=hexafluoroacetylacetonate; and NITPh-3-Br-4-OMe=2-3´-Br-4´-methoxyphenyl-4,4,5,5 -tetramethylimidazoline-1-oxyl-3-oxide). The single-crystal structures analyses show that these complexes have similar mononuclear tri-spin structures, in which central Ln(III) ions are all eight coordinated by three hfac molecules and two NITPh-3-Br-4-OMe radicals. The variable-temperature magnetic susceptibility studies reveal the antiferromagnetic interactions between the paramagnetic ions (Ln(III) and radicals) in complexes 1, 2, 3 and 5 and ferromagnetic interaction in complex 4. The luminescence characterizations of complexes Eu(1), Tb(3) and Dy(4) are also studied in this paper. - Graphical abstract: Using a novel halogen phenyl-substituted nitronyl-nitroxide radical, we obtained and characterized five isostructural lanthanide mononuclear tri-spin compounds. Highlights: • A new halogen phenyl-substituted nitronyl-nitroxide radical was designed. • Five new Ln(III) radical complexes have been synthesized and characterized. • The reasonable evaluation the magnetic interactions between Ln(III) ions and radical is meaningful. • These complexes show good luminescent properties.

  15. Flow injection fluorometric determination of ascorbic acid using perylenebisimide-linked nitroxide.

    PubMed

    Maki, Tomoharu; Soh, Nobuaki; Nakano, Koji; Imato, Toshihiko

    2011-09-30

    A simple and sensitive flow injection fluorometric method for the determination of ascorbic acid is described. Perylenebisimide-linked nitroxide (PBILN) is used as a fluorescent reagent, which permits the selective determination of ascorbic acid. The fluorescence of the perylenebisimide moiety in PBILN is quenched by the nitroxide moiety, which is linked to the perylenebisimide. When a stream of a solution of ascorbic acid is merged with a stream of PBILN, the ascorbic acid reacts with the nitroxide moiety of PBILN to form hydroxylamine, and the fluorescence properties of the perylenebisimide moiety are recovered. As a result, a peak-shaped fluorescence signal is produced, which can be observed by a fluorescence detector located downstream. Under optimized conditions, a good linear relationship between the concentration of ascorbic acid and peak height in the concentration range from 0.5 to 10 μmol L(-1) was found and the detection limit (S/N=3) was 0.28 μmol L(-1). The relative standard deviation for the determination of 4.0 μmol L(-1) ascorbic acid samples was 1.0% (n=5). The proposed method was applied to the determination of ascorbic acid in several soft drink beverages and the analytical results were in good agreement with those obtained using a conventional method.

  16. Simultaneous 280 MHz EPR imaging of rat organs during nitroxide free radical clearance.

    PubMed Central

    Alecci, M; Ferrari, M; Quaresima, V; Sotgiu, A; Ursini, C L

    1994-01-01

    A radio frequency (RF) (280 MHz) electron paramagnetic resonance (EPR) spectroscopy and imaging apparatus has been used to localize a pyrrolidine nitroxide free radical in the rat abdomen and thorax. The nitroxide 2,2.5.5,-tetramethylpyrrolidine-1-oxyl-3- carboxylic acid (PCA) had a whole body monoexponential decay with half-life of 13.3 +/- 0.7 (n = 4), 19.4 +/- 0.2 (n = 3), and 23 +/- 2 (n = 6) min for 1, 2, and 3 mmol/kg PCA, respectively. Up to seven one-dimensional longitudinal projections were collected on six rats in the presence of a 8 mT/m field gradient. With an injection dose of 3 mmol/kg, PCA half-lives were 19 +/- 1, 17 +/- 2, and 22 +/- 2 min (n = 6) in the lower abdomen, in the liver, and in the thorax, respectively. Thorax half-life was significantly longer than liver half-life. Sequential two-dimensional images of PCA distribution in a plane longitudinal to the rat body were obtained from eight spectra in the presence of a gradient of 12 mT/m (acquisition time 5 min; spatial resolution 8 mm). After 7 min, the nitroxide was detectable in the left side of the thorax area, but it was mostly localized in the liver. PCA was more uniformly distributed in the image collected after 17 min. Images FIGURE 4 PMID:7811942

  17. Discriminative EPR detection of NO and HNO by encapsulated nitronyl nitroxides

    PubMed Central

    Bobko, Andrey A.; Ivanov, Alexander; Khramtsov, Valery V.

    2014-01-01

    Nitric oxide, •NO, is one of the most important molecules in the biochemistry of living organisms. By contrast, nitroxyl, NO−, one-electron reduced analog of •NO which exists at physiological conditions in its protonated form, HNO, has been relatively overlooked. Recent data shows that HNO might be produced endogenously and display unique biological effects. However, there is a lack of specific and quantitative methods of detection of endogenous HNO production. Here we present a new method for discriminative •NO and HNO detection by nitronyl nitroxides (NNs) using electron paramagnetic resonance (EPR). It was found that NNs react with •NO and HNO with similar rate constants of about 104 M−1s−1 but yield different products: imino nitroxides and the hydroxylamine of imino nitroxides, correspondingly. An EPR approach for discriminative •NO and HNO detection using liposome-encapsulated NNs was developed. The membrane barrier of liposomes protects NNs against reduction in biological systems while is permeable to both analytes, •NO and HNO. The sensitivity of this approach for the detection of the rates of •NO/HNO generation is about 1 nM/s. The application of encapsulated NNs for real-time discriminative •NO/HNO detection might become a valuable tool in nitric oxide related studies. PMID:23136998

  18. Oxygen concentration dependence of lipid peroxidation and lipid-derived radical generation: application of profluorescent nitroxide switch.

    PubMed

    Mito, Fumiya; Kitagawa, Kana; Yamasaki, Toshihide; Shirahama, Chisato; Oishi, Taketoshi; Ito, Yuko; Yamato, Mayumi; Yamada, Ken-Ichi

    2011-09-01

    Lipid-derived radicals and peroxides are involved in the pathogenesis of oxidative stress diseases and, although lipid peroxide production is a required reaction between a lipid radical and molecular oxygen, a useful lipid radical detection method has remained tentative. Also, the effect of oxygen concentration on lipid peroxide production must be considered because of the hypoxic conditions in cancer and ischemic regions. In this study, the focus was on nitroxide reactivity, which allows spin trapping with carbon-centred radicals via radical-radical reactions and fluorophore quenching through interactions with nitroxide's unpaired electron. Thus, the aim here was to demonstrate a useful detection method for lipid-derived radicals as well as to clarify the effects of oxygen concentration on lipid peroxide production using profluorescent nitroxide. This latter compound reacted with lipid-derived radicals in a manner inversely dependent on oxygen concentration, resulting in fluorescence due to alkoxyamine formation and, conversely, lipid peroxide concentrations decreased with lower oxygen in the reaction system. Furthermore, nitroxide inhibited lipid peroxide production and stopped oxygen consumption in the same solution. These results suggested that the novel application of profluorescent nitroxide could directly and sensitively detect lipid-derived radicals and that radical and peroxide production were dependent on oxygen concentration.

  19. Understanding the fundamentals of redox mediators in Li-O2 batteries: a case study on nitroxides.

    PubMed

    Bergner, Benjamin J; Hofmann, Christine; Schürmann, Adrian; Schröder, Daniel; Peppler, Klaus; Schreiner, Peter R; Janek, Jürgen

    2015-12-21

    The development of aprotic lithium-oxygen (Li-O2) batteries suffers from high charging overvoltages. Dissolved redox mediators, like nitroxides, providing increased energy efficiency and longer lifetime are promising tools to overcome this challenge. Since this auspicious concept is still in its infancy, the underlying chemical reactions as well as the impact of the different (electro)chemical parameters are poorly understood. Herein, we derive an electrochemical model for the charging reactions, which is validated by potentiostatic measurements. The model elucidates the impact of the major factors including basic cell parameters and the chemical properties of the redox mediator. The model is applied to the promising class of nitroxides, which is systematically investigated by using derivatives of TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxy), AZADO (2-azaadamantane-N-oxyl), and an azaphenalene based nitroxide. The nitroxides are electrochemically characterized by cyclic voltammetry and their performance as redox mediators is studied in Li-O2 batteries with an ether-based electrolyte. Based on the presented model, the charging profiles of the different nitroxide redox mediators are correlated with their molecular structures.

  20. Ferromagnetic interactions between imino nitroxides through diamagnetic metal ions: Crystal structures, magnetism, and electronic properties of [M{sup I}(imino nitroxide){sub 2}](PF{sub 6}) (M = Cu{sup I} and Ag{sup I})

    SciTech Connect

    Oshio, Hiroki; Watanabe, Takashi; Ohto, Akihiro

    1997-07-02

    Ferromagnetic interactions between imino nitroxides through Diamagnetic Metal Ions: Crystal Structures, Magnetism, and Electronic Properties of [M{sup I}(imino nitroxide){sub 2}](PF{sub 6}) (M = Cu{sup I} and Ag{sup I}) Cu(I) and Ag(I) complexes with imino nitroxides, [Cu{sup I}-(immepy){sub 2}](PF{sub 6}) and [Ag{sup I}(impy){sub 2}](PF{sub 6}), were structurally and magnetically characterized. Magnetic susceptibility and EPR measurements revealed that the magnetic interaction in the Cu(I) complex is ferromagnetic with a J value of 55.1(6) cm{sup -1} (g = 2.0 and H = -JS{sub 1}{center_dot}S{sub 2}), while the Ag(I) complex shows a very weak ferromagnetic interaction. The magnetic behaviors were discussed in terms of orthogonality of magnetic orbitals and metal to ligand charge-transfer interactions.

  1. The nitroxide TEMPO is an efficient scavenger of protein radicals: cellular and kinetic studies.

    PubMed

    Pattison, David I; Lam, Magdalena; Shinde, Sujata S; Anderson, Robert F; Davies, Michael J

    2012-11-01

    Protein oxidation occurs during multiple human pathologies, and protein radicals are known to induce damage to other cell components. Such damage may be modulated by agents that scavenge protein radicals. In this study, the potential protective reactions of the nitroxide TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxyl radical) against Tyr- and Trp-derived radicals (TyrO./TrpN.) have been investigated. Pretreatment of macrophage cells with TEMPO provided protection against photo-oxidation-induced loss of cell viability and Tyr oxidation, with the nitroxide more effective than the hydroxylamine or parent amine. Pulse radiolysis was employed to determine rate constants, k, for the reaction of TEMPO with TyrO. and TrpN. generated on N-Ac-Tyr-amide and N-Ac-Trp-amide, with values of k~10(8) and 7×10(6)M(-1)s(-1), respectively, determined. Analogous studies with lysozyme, chymotrypsin, and pepsin yielded k for TEMPO reacting with TrpN. ranging from 1.5×10(7) (lysozyme) to 1.1×10(8) (pepsin)M(-1)s(-1). Pepsin-derived TyrO. reacted with TEMPO with k~4×10(7)M(-1)s(-1); analogous reactions for lysozyme and chymotrypsin TyrO. were much slower. These data indicate that TEMPO can inhibit secondary reactions of both TyrO. and TrpN., though this is protein dependent. Such protein radical scavenging may contribute to the positive biological effects of nitroxides.

  2. A nucleotide-independent cyclic nitroxide label for monitoring segmental motions in nucleic acids.

    PubMed

    Nguyen, Phuong H; Popova, Anna M; Hideg, Kálmán; Qin, Peter Z

    2015-01-01

    Spin labels, which are chemically stable radicals attached at specific sites of a bio-molecule, enable investigations on structure and dynamics of proteins and nucleic acids using techniques such as site-directed spin labeling and paramagnetic NMR. Among spin labels developed, the class of rigid labels have limited or no independent motions between the radical bearing moiety and the target, and afford a number of advantages in measuring distances and monitoring local dynamics within the parent bio-molecule. However, a general method for attaching a rigid label to nucleic acids in a nucleotide-independent manner has not been reported. We developed an approach for installing a nearly rigid nitroxide spin label, designated as R5c, at a specific site of the nucleic acid backbone in a nucleotide-independent manner. The method uses a post-synthesis approach to covalently attach the nitroxide moiety in a cyclic fashion to phosphorothioate groups introduced at two consecutive nucleotides of the target strand. R5c-labeled nucleic acids are capable of pairing with their respective complementary strands, and the cyclic nature of R5c attachment significantly reduced independence motions of the label with respect to the parent duplex, although it may cause distortion of the local environment at the site of labeling. R5c yields enhanced sensitivity to the collective motions of the duplex, as demonstrated by its capability to reveal changes in collective motions of the substrate recognition duplex of the 120-kDa Tetrahymena group I ribozyme, which elude detection by a flexible label. The cyclic R5c nitroxide can be efficiently attached to a target nucleic acid site using a post-synthetic coupling approach conducted under mild biochemical conditions, and serves as a viable label for experimental investigation of segmental motions in nucleic acids, including large folded RNAs.

  3. The impact of carboxy nitroxide antioxidants on irradiated ataxia telangiectasia cells.

    PubMed

    Hosokawa, Kazuyuki; Chen, Philip; Lavin, F Martin; Bottle, E Steven

    2004-10-01

    Three water-soluble carboxy nitroxide antioxidants, 5-carboxy-1,1,3,3-tetramethylisoindolin-2-yloxyl, 4-carboxy-2,2,6,6-tetramethylpiperidin-1-yloxyl, and 3-carboxy-2,2,5,5-tetramethylpyrrolidin-1-yloxyl, show significant impact on the postirradiation survival rates of ataxia telangiectasia (A-T) cells compared to normal cells, an assay which represents a model for understanding the impact of ROS damage on the A-T phenotype. The effects of these antioxidants are much more significant than those of vitamin E or Trolox (a water-soluble vitamin E analog), studied using the same cell survival model.

  4. Cyclic nitroxide radicals attenuate inflammation and Hyper-responsiveness in a mouse model of allergic asthma.

    PubMed

    Assayag, Miri; Goldstein, Sara; Samuni, Amram; Berkman, Neville

    2015-10-01

    The effects of stable cyclic nitroxide radicals have been extensively investigated both in vivo and in vitro demonstrating anti-inflammatory, radioprotective, anti-mutagenic, age-retardant, hypotensive, anti-cancer and anti-teratogenic activities. Yet, these stable radicals have not been evaluated in asthma and other airway inflammatory disorders. The present study investigated the effect of 4-hydroxy-2,2,6,6-tetramethyl-piperidine-N-oxyl (TPL) and 3-carbamoyl-proxyl (3-CP) in a mouse model of ovalbumin (OVA)-induced allergic asthma. Both 3-CP and TPL were non-toxic when administered either orally (1% w/w nitroxide-containing chow) or via intraperitoneal (IP) injection (∼300 mg/kg). Feeding the mice orally demonstrated that 3-CP was more effective than TPL in reducing inflammatory cell recruitment into the airway and in suppressing airway hyper-responsiveness (AHR) in OVA-challenged mice. To characterize the optimal time-window of intervention and mode of drug administration, 3-CP was given orally during allergen sensitization, during allergen challenge or during both sensitization and challenge stages, and via IP injection or intranasal instillation for 3 days during the challenge period. 3-CP given via all modes of delivery markedly inhibited OVA-induced airway inflammation, expression of cytokines, AHR and protein nitration of the lung tissue. Oral administration during the entire experiment was the most efficient delivery of 3-CP and was more effective than dexamethasone a potent corticosteroid used for asthma treatment. Under a similar administration regimen (IP injection before the OVA challenge), the effect of 3-CP was similar to that of dexamethasone and even greater on AHR and protein nitration. The protective effect of the nitroxides, which preferentially react with free radicals, in suppressing the increase of main asthmatic inflammatory markers substantiate the key role played by reactive oxygen and nitrogen species in the molecular mechanism of

  5. High performance liquid chromatography and electron spin resonance studies of some sugar-nitroxide solutions

    SciTech Connect

    Angel, J.P.; Thiery, C.; Battesti, C.; Vincent, P.; Raffi, J.

    1985-01-01

    Radicals induced by gamma irradiation of alpha-D-glucose, 1-0-methyl-alpha-D-glucose and maltose, in the solid state, have been studied by the spin-trapping method. High performance liquid chromatography of sugar-nitroxide solutions, combined with electron spin resonance analysis, revealed nine, eight and twelve discernible radical species, the majority of them being indiscernible by the direct spin-trapping method. Tentative correlations and assignments of chemical structures are discussed. 9 references, 4 figures, 3 tables.

  6. Electron dipole-dipole ESEEM in field-step ELDOR of nitroxide biradicals.

    PubMed

    Kulik, L V; Grishin, Yu A; Dzuba, S A; Grigoryev, I A; Klyatskaya, S V; Vasilevsky, S F; Tsvetkov, Yu D

    2002-07-01

    The use of a rapid stepping of the magnetic field for investigation of electron dipole-dipole ESEEM in pulsed X-band ELDOR is described. The magnetic field jump, synchronized with a microwave pumping pulse, is positioned between the second and the third pulses of the stimulated echo pulse sequence. This echo is measured as a function of the delay between the first and the second pulses. The data are analyzed for a Fourier transform resulting in a Pake resonance pattern. To remove the electron-nuclear contributions to ESEEM, time traces with pumping were divided by those without. This resulted in complete elimination of electron-nuclear contributions, which is seen from the absence of peaks at nuclear frequencies and the similarity of results for protonated and deuterated solvents. For increasing the electron-electron modulation depth, a scanning of the magnetic field during the microwave pumping is proposed. The interspin distances and their distribution are determined for two long-chained (ca. 2 nm) nitroxide biradicals in glassy toluene and in frozen nematic liquid crystal 4-cyano-4'-pentyl-biphenyl. For the latter solvent, the alignment of the axis connecting two nitroxides in biradicals is quantitatively analyzed.

  7. Electron Dipole-Dipole ESEEM in Field-Step ELDOR of Nitroxide Biradicals

    NASA Astrophysics Data System (ADS)

    Kulik, L. V.; Grishin, Yu. A.; Dzuba, S. A.; Grigoryev, I. A.; Klyatskaya, S. V.; Vasilevsky, S. F.; Tsvetkov, Yu. D.

    2002-07-01

    The use of a rapid stepping of the magnetic field for investigation of electron dipole-dipole ESEEM in pulsed X-band ELDOR is described. The magnetic field jump, synchronized with a microwave pumping pulse, is positioned between the second and the third pulses of the stimulated echo pulse sequence. This echo is measured as a function of the delay between the first and the second pulses. The data are analyzed for a Fourier transform resulting in a Pake resonance pattern. To remove the electron-nuclear contributions to ESEEM, time traces with pumping were divided by those without. This resulted in complete elimination of electron-nuclear contributions, which is seen from the absence of peaks at nuclear frequencies and the similarity of results for protonated and deuterated solvents. For increasing the electron-electron modulation depth, a scanning of the magnetic field during the microwave pumping is proposed. The interspin distances and their distribution are determined for two long-chained (ca. 2 nm) nitroxide biradicals in glassy toluene and in frozen nematic liquid crystal 4-cyano-4'-pentyl-biphenyl. For the latter solvent, the alignment of the axis connecting two nitroxides in biradicals is quantitatively analyzed.

  8. Fluorescent dye-labelled polymer synthesis by nitroxide mediated radical polymerization

    NASA Astrophysics Data System (ADS)

    Kollár, Jozef; Chmela, Štefan; Hrčková, Ľudmila; Hrdlovič, Pavol

    2012-07-01

    New applications of polymers at advanced technologies demand increased requirements on their properties. These properties are influenced by molecular as well as supramolecular structure. Controlled radical polymerization mediated by stable nitroxides (NMP) or substituted alkoxyamines offers simple method for preparation of polymers with programmable structure of macromolecules which possess remarkable better physical as well as chemical properties. They can be used as a macro initiators for the synthesis of block copolymers. At the present time it has been generally accepted that the extent of "livingness" is high for all conversions [1-4]. To verify this statement a series of fluorescent dye-labelled regulators has been synthesized, spectrally characterized and used as the mediators of styrene and n-butyl acrylate polymerization. Direct quantification of dormant species concentration (extent of livingness) and calculation of molar mass of marked polymers was performed by absorption and/or emission spectroscopy. Controlled radical polymerization mediated by stable nitroxides bearing fluorescence mark represents unconventional approach for monitoring and evaluation of mechanism and kinetics of polymerization process. Results indicate that the extent of livingness is strongly influenced by conversion as well as mediator concentration. There is a clear tendency toward to decreasing amount of dormant species with increasing monomer conversion. Moreover, lower mediator concentration decreases livingness of polymerization process.

  9. Nitroxide/substrate weak hydrogen bonding: attitude and dynamics of collisions in solution.

    PubMed

    Russ, Jennifer L; Gu, Juan; Tsai, Kun-Hsiang; Glass, Tom; Duchamp, James C; Dorn, Harry C

    2007-06-06

    The study of intermolecular collisions and bonding interactions in solutions is of critical importance in understanding and predicting solute/solvent properties. Previous work has established that stable paramagnetic nitroxide molecules are excellent probes of intermolecular interactions for hydrogen bonding in polar solvents. In this study, 1H, 2H, 13C, 15N NMR and liquid/liquid intermolecular transfer dynamic nuclear polarization (L2IT DNP) results are obtained for the paramagnetic probe molecule, TEMPO, interacting with the common aprotic and protic polar solvents, CH3CN and CH3CONH2, yielding a profile of both dipolar and scalar interactions. A significant scalar contact hyperfine is observed for the N-O...H-C interaction (13CH3 hyperfine, a/h=0.66 MHz) in the CH3CN/TEMPO system, whereas the N-O...H-C and N-O...H-N interactions for the TEMPO/CH3CONH2 system yield 13CH3 and 15N hyperfine couplings of a/h=0.16 and -0.50 MHz, respectively. The distance and attitude of the scalar interaction for the nitroxide hydrogen bonding at the methyl group in CH3CN and the amino group in CH3CONH2 are computed using density functional theory (DFT), yielding good agreement with the experimental results. These results show that the hyperfine coupling provides a sensitive probe of weak hydrogen-bonding interactions in solution.

  10. Controlled Redox Chemistry at Cerium within a Tripodal Nitroxide Ligand Framework

    DOE PAGES

    Bogart, Justin A.; Lippincott, Connor A.; Carroll, Patrick J.; ...

    2015-10-27

    Ligand reorganization has been shown to have a profound effect on the outcome of cerium redox chemistry. Through the use of a tethered, tripodal, trianionic nitroxide ligand, [((2-tBuNOH)C 6 H 4 CH 2 ) 3 N] 3- (TriNO x 3- ), controlled redox chemistry at cerium was accomplished, and typically reactive complexes of tetravalent cerium were isolated. These included rare cationic complexes [Ce(TriNO x )thf][BAr F 4 ], in which Ar F =3,5-(CF 3 ) 2 -C 6 H 3 , and [Ce(TriNO x )py][OTf] . A rare complete Ce-halide series, Ce(TriNO x )X, in which X=F - , Clmore » - , Br - , I - , was also synthesized. We explored the solution chemistry of these complexes through detailed solution-phase electrochemistry and 1 H NMR experiments and showed a unique shift in the ratio of species with inner- and outer-sphere anions with size of the anionic X - group. DFT calculations on the series of calculations corroborated the experimental findings. Also, the use of a bulky and strongly donating tethered tripodal nitroxide ligand allowed the controlled redox chemistry at cerium. As a result, rare examples of cationic Ce IV complexes were synthesized and fully characterized. The full Ce-halide series supported by the tripodal ligand framework is also reported (see scheme).« less

  11. Relaxation-based distance measurements between a nitroxide and a lanthanide spin label

    NASA Astrophysics Data System (ADS)

    Jäger, H.; Koch, A.; Maus, V.; Spiess, H. W.; Jeschke, G.

    2008-10-01

    Distance measurements by electron paramagnetic resonance techniques between labels attached to biomacromolecules provide structural information on systems that cannot be crystallized or are too large to be characterized by NMR methods. However, existing techniques are limited in their distance range and sensitivity. It is anticipated by theoretical considerations that these limits could be extended by measuring the enhancement of longitudinal relaxation of a nitroxide label due to a lanthanide complex label at cryogenic temperatures. The relaxivity of the dysprosium complex with the macrocyclic ligand DOTA can be determined without direct measurements of longitudinal relaxation rates of the lanthanide and without recourse to model compounds with well defined distance by analyzing the dependence of relaxation enhancement on either temperature or concentration in homogeneous glassy frozen solutions. Relaxivities determined by the two calibration techniques are in satisfying agreement with each other. Error sources for both techniques are examined. A distance of about 2.7 nm is measured in a model compound of the type nitroxide-spacer-lanthanide complex and is found in good agreement with the distance in a modeled structure. Theoretical considerations suggest that an increase of the upper distance limit requires measurements at lower fields and temperatures.

  12. Increased Nitroxidative Stress Promotes Mitochondrial Dysfunction in Alcoholic and Nonalcoholic Fatty Liver Disease

    PubMed Central

    Song, Byoung-Joon; Abdelmegeed, Mohamed A.; Henderson, Lauren E.; Yoo, Seong-Ho; Wan, Jie; Purohit, Vishnudutt; Hardwick, James P.; Moon, Kwan-Hoon

    2013-01-01

    Increased nitroxidative stress causes mitochondrial dysfunctions through oxidative modifications of mitochondrial DNA, lipids, and proteins. Persistent mitochondrial dysfunction sensitizes the target cells/organs to other pathological risk factors and thus ultimately contributes to the development of more severe disease states in alcoholic and nonalcoholic fatty liver disease. The incidences of nonalcoholic fatty liver disease continuously increase due to high prevalence of metabolic syndrome including hyperlipidemia, hypercholesterolemia, obesity, insulin resistance, and diabetes. Many mitochondrial proteins including the enzymes involved in fat oxidation and energy supply could be oxidatively modified (including S-nitrosylation/nitration) under increased nitroxidative stress and thus inactivated, leading to increased fat accumulation and ATP depletion. To demonstrate the underlying mechanism(s) of mitochondrial dysfunction, we employed a redox proteomics approach using biotin-N-maleimide (biotin-NM) as a sensitive biotin-switch probe to identify oxidized Cys residues of mitochondrial proteins in the experimental models of alcoholic and acute liver disease. The aims of this paper are to briefly describe the mechanisms, functional consequences, and detection methods of mitochondrial dysfunction. We also describe advantages and limitations of the Cys-targeted redox proteomics method with alternative approaches. Finally, we discuss various applications of this method in studying oxidatively modified mitochondrial proteins in extrahepatic tissues or different subcellular organelles and translational research. PMID:23691267

  13. Synthesis, crystal structure, superoxide scavenging activity, anticancer and docking studies of novel adamantyl nitroxide derivatives

    NASA Astrophysics Data System (ADS)

    Zhu, Xiao-he; Sun, Jin; Wang, Shan; Bu, Wei; Yao, Min-na; Gao, Kai; Song, Ying; Zhao, Jin-yi; Lu, Cheng-tao; Zhang, En-hu; Yang, Zhi-fu; Wen, Ai-dong

    2016-03-01

    A novel adamantyl nitroxide derivatives has been synthesized and characterized by IR, ESI-MS and elemental analysis. Quantum chemical calculations have also been performed to calculate the molecular geometry using density functional theory (B3LYP) with the 6-31G (d,p) basis set. The calculated results showed that the optimized geometry can well reproduce the crystal structure. The antioxidant and antiproliferative activity were evaluated by superoxide (NBT) and MTT assay. The adamantyl nitroxide derivatives exhibited stronger scavenging ability towards O2· - radicals when compared to Vitamin C, and demonstrated a remarked anticancer activity against all the tested cell lines, especially Bel-7404 cells with IC50 of 43.3 μM, compared to the positive control Sorafenib (IC50 = 92.0 μM). The results of molecular docking within EGFR using AutoDock confirmed that the titled compound favorably fitted into the ATP binding site of EGFR and would be a potential anticancer agent.

  14. Interactions of Nitroxide-Conjugated and Non-Conjugated Glycodendrimers with Normal and Cancer Cells and Biocompatibility Studies.

    PubMed

    Andreozzi, Elisa; Antonelli, Antonella; Cangiotti, Michela; Canonico, Barbara; Sfara, Carla; Pianetti, Anna; Bruscolini, Francesca; Sahre, Karin; Appelhans, Dietmar; Papa, Stefano; Ottaviani, Maria Francesca

    2017-02-15

    Poly(propyleneimine) glycodendrimers fully modified with maltose units were administered to different cancer cell lines and their effect on cell viability was evaluated by using MTS assay and flow cytometry. The mechanism of dendrimer-cell interactions was investigated by the electron paramagnetic resonance (EPR) technique by using a new nitroxide-conjugated glycodendrimer. The nitroxide groups did not modify both the biological properties (cell viability and apoptosis degree) of the dendrimers in the presence of the cells and the dendrimer-cell interactions. Since this class of dendrimers is already known to be biocompatible for human healthy cells, noncancer cells such as human peripheral blood mononuclear cells (PBMCs) and macrophages were also treated with the glycodendrimer, and EPR spectra of the nitroxide-conjugated glycodendrimer were compared for cancer and noncancer cells. It was found that this dendrimer selectively affects the cell viability of tumor cells, while, surprisingly, PBMC proliferation is induced. Moreover, H-bond-active glycodendrimer-cell interactions were different for the different cancer cell lines and noncancer cells. The nitroxide-conjugated glycodendrimer was able to interact with the cell membrane and eventually cross it, getting in contact with cytosol antioxidants. This study helps to clarify the potential anticancer effect of this class of dendrimers opening to future applications of these macromolecules as new antitumor agents.

  15. Interaction of poly(ethylene oxide) with the sodium dodecyl sulfate micelle interface studied with nitroxide spin probes

    SciTech Connect

    Kang, Y.S.; Kevan, L. )

    1994-08-04

    Electron spin resonance (ESR) line widths of 5-, 7-, 12-, and 16-doxylstearic acid (x-DSA) and tempo nitroxides versus the concentration of poly(ethylene oxide) (PEO) in sodium dodecyl sulfate (SDS) micelles show different trends. The ESR line widths of 5-, 7-, and 16-DSA increase with increasing concentration of PEO, which is interpreted as due to increasing viscosity in the environment of the nitroxide spin probe. The tempo and 12-DSA line widths were independent of the concentration of PEO. The line width showed the highest value for 5-DSA and the lowest value of tempo. The line width of x-DSA decreases from 5-DSA to a minimum value for 12-DSA and then increases somewhat for 16-DSA. This is interpreted as bending of the alkyl chain to provide different locations for the nitroxide moiety relative to the micelle interface. The relative distances of the nitroxide moiety of [chi]-DSA from deuterated water at the SDS micelle interface was measured by deuterium electron spin echo modulation. The distances increased from 5-DSA to 12-DSA and then decreased for 16-DSA. The interpretation of the DSR line width trend is supported by the deuterium modulation depth trend. 28 refs., 5 figs., 2 tabs.

  16. Distance determination between low-spin ferric haem and nitroxide spin label using DEER: the neuroglobin case

    NASA Astrophysics Data System (ADS)

    Ezhevskaya, M.; Bordignon, E.; Polyhach, Y.; Moens, L.; Dewilde, S.; Jeschke, G.; Van Doorslaer, S.

    2013-10-01

    This work demonstrates for the first time the feasibility of using double electron-electron resonance (DEER) to determine the inter-spin distance between nitroxide spin labels and low-spin (S = 1/2) ferric haem centres. For these means, two human neuroglobin variants were spin labelled leading to singly labelled haem proteins with the nitroxide label on one of the natural Cys residues (Cys55 or Cys120). Room-temperature electron paramagnetic resonance was used to characterise the mobility of the nitroxide labels and X- and Q-band DEER experiments were performed to detect nitroxide-haem distances. Effects of residual nuclear modulations in the DEER traces were carefully evaluated. The DEER-derived distances were compared with theoretical predictions from an X-ray diffraction structure of human neuroglobin using a rotamer library approach as well as with distance information obtained from electron relaxation measurements. The structural biological implications of the spin-labelled side chains' dynamics and of the obtained distances are also discussed.

  17. Relaxation Times and Line Widths of Isotopically-Substituted Nitroxides in Aqueous Solution at X-band

    PubMed Central

    Biller, Joshua R.; Meyer, Virginia; Elajaili, Hanan; Rosen, Gerald M.; Kao, Joseph P.Y.; Eaton, Sandra S.; Eatona, Gareth R.

    2011-01-01

    Optimization of nitroxides as probes for EPR imaging requires detailed understanding of spectral properties. Spin lattice relaxation times, spin packet line widths, nuclear hyperfine splitting, and overall lineshapes were characterized for six low molecular weight nitroxides in dilute deoxygenated aqueous solution at X-band. The nitroxides included 6-member, unsaturated 5-member, or saturated 5-member rings, most of which were isotopically labeled. The spectra are near the fast tumbling limit with T1 ~ T2 in the range of 0.50 to 1.1 μs at ambient temperature. Both spin-lattice relaxation T1 and spin-spin relaxation T2 are longer for 15N- than for 14N-nitroxides. The dominant contributions to T1 are modulation of nitrogen hyperfine anisotropy and spin rotation. Dependence of T1 on nitrogen nuclear spin state mI was observed for both 14N and 15N. Unresolved hydrogen/deuterium hyperfine couplings dominate overall line widths. Lineshapes were simulated by including all nuclear hyperfine couplings and spin packet line widths that agreed with values obtained by electron spin echo. Line widths and relaxation times are predicted to be about the same at 250 MHz as at X-band. PMID:21843961

  18. Mitochondria-targeted catalase reverts the neurotoxicity of hSOD1G⁹³A astrocytes without extending the survival of ALS-linked mutant hSOD1 mice.

    PubMed

    Pehar, Mariana; Beeson, Gyda; Beeson, Craig C; Johnson, Jeffrey A; Vargas, Marcelo R

    2014-01-01

    Dominant mutations in the Cu/Zn-superoxide dismutase (SOD1) cause familial forms of amyotrophic lateral sclerosis (ALS), a fatal disorder characterized by the progressive loss of motor neurons. The molecular mechanism underlying the toxic gain-of-function of mutant hSOD1s remains uncertain. Several lines of evidence suggest that toxicity to motor neurons requires damage to non-neuronal cells. In line with this observation, primary astrocytes isolated from mutant hSOD1 over-expressing rodents induce motor neuron death in co-culture. Mitochondrial alterations have been documented in both neuronal and glial cells from ALS patients as well as in ALS-animal models. In addition, mitochondrial dysfunction and increased oxidative stress have been linked to the toxicity of mutant hSOD1 in astrocytes and neurons. In mutant SOD1-linked ALS, mitochondrial alterations may be partially due to the increased association of mutant SOD1 with the outer membrane and intermembrane space of the mitochondria, where it can affect several critical aspects of mitochondrial function. We have previously shown that decreasing glutathione levels, which is crucial for peroxide detoxification in the mitochondria, significantly accelerates motor neuron death in hSOD1G93A mice. Here we employed a catalase targeted to the mitochondria to investigate the effect of increased mitochondrial peroxide detoxification capacity in models of mutant hSOD1-mediated motor neuron death. The over-expression of mitochondria-targeted catalase improved mitochondrial antioxidant defenses and mitochondrial function in hSOD1G93A astrocyte cultures. It also reverted the toxicity of hSOD1G93A-expressing astrocytes towards co-cultured motor neurons, however ALS-animals did not develop the disease later or survive longer. Hence, while increased oxidative stress and mitochondrial dysfunction have been extensively documented in ALS, these results suggest that preventing peroxide-mediated mitochondrial damage alone is not

  19. Mitochondria-targeted plastoquinone derivatives as tools to interrupt execution of the aging program. 4. Age-related eye disease. SkQ1 returns vision to blind animals.

    PubMed

    Neroev, V V; Archipova, M M; Bakeeva, L E; Fursova, A Zh; Grigorian, E N; Grishanova, A Yu; Iomdina, E N; Ivashchenko, Zh N; Katargina, L A; Khoroshilova-Maslova, I P; Kilina, O V; Kolosova, N G; Kopenkin, E P; Korshunov, S S; Kovaleva, N A; Novikova, Yu P; Philippov, P P; Pilipenko, D I; Robustova, O V; Saprunova, V B; Senin, I I; Skulachev, M V; Sotnikova, L F; Stefanova, N A; Tikhomirova, N K; Tsapenko, I V; Shchipanova, A I; Zinovkin, R A; Skulachev, V P

    2008-12-01

    Mitochondria-targeted cationic plastoquinone derivative SkQ1 (10-(6'-plastoquinonyl) decyltriphenylphosphonium) has been investigated as a potential tool for treating a number of ROS-related ocular diseases. In OXYS rats suffering from a ROS-induced progeria, very small amounts of SkQ1 (50 nmol/kg per day) added to food were found to prevent development of age-induced cataract and retinopathies of the eye, lipid peroxidation and protein carbonylation in skeletal muscles, as well as a decrease in bone mineralization. Instillation of drops of 250 nM SkQ1 reversed cataract and retinopathies in 3-12-month-old (but not in 24-month-old) OXYS rats. In rabbits, experimental uveitis and glaucoma were induced by immunization with arrestin and injections of hydroxypropyl methyl cellulose to the eye anterior sector, respectively. Uveitis was found to be prevented or reversed by instillation of 250 nM SkQ1 drops (four drops per day). Development of glaucoma was retarded by drops of 5 microM SkQ1 (one drop daily). SkQ1 was tested in veterinarian practice. A totally of 271 animals (dogs, cats, and horses) suffering from retinopathies, uveitis, conjunctivitis, and cornea diseases were treated with drops of 250 nM SkQ1. In 242 cases, positive therapeutic effect was obvious. Among animals suffering from retinopathies, 89 were blind. In 67 cases, vision returned after SkQ1 treatment. In ex vivo studies of cultivated posterior retina sector, it was found that 20 nM SkQ1 strongly decreased macrophagal transformation of the retinal pigmented epithelial cells, an effect which might explain some of the above SkQ1 activities. It is concluded that low concentrations of SkQ1 are promising in treating retinopathies, cataract, uveitis, glaucoma, and some other ocular diseases.

  20. Superoxide activates uncoupling proteins by generating carbon-centered radicals and initiating lipid peroxidation: studies using a mitochondria-targeted spin trap derived from alpha-phenyl-N-tert-butylnitrone.

    PubMed

    Murphy, Michael P; Echtay, Karim S; Blaikie, Frances H; Asin-Cayuela, Jordi; Cocheme, Helena M; Green, Katherine; Buckingham, Julie A; Taylor, Ellen R; Hurrell, Fiona; Hughes, Gillian; Miwa, Satomi; Cooper, Christopher E; Svistunenko, Dimitri A; Smith, Robin A J; Brand, Martin D

    2003-12-05

    Although the physiological role of uncoupling proteins (UCPs) 2 and 3 is uncertain, their activation by superoxide and by lipid peroxidation products suggest that UCPs are central to the mitochondrial response to reactive oxygen species. We examined whether superoxide and lipid peroxidation products such as 4-hydroxy-2-trans-nonenal act independently to activate UCPs, or if they share a common pathway, perhaps by superoxide exposure leading to the formation of lipid peroxidation products. This possibility can be tested by blocking the putative reactive oxygen species cascade with selective antioxidants and then reactivating UCPs with distal cascade components. We synthesized a mitochondria-targeted derivative of the spin trap alpha-phenyl-N-tert-butylnitrone, which reacts rapidly with carbon-centered radicals but is unreactive with superoxide and lipid peroxidation products. [4-[4-[[(1,1-Dimethylethyl)-oxidoimino]methyl]phenoxy]butyl]triphenylphosphonium bromide (MitoPBN) prevented the activation of UCPs by superoxide but did not block activation by hydroxynonenal. This was not due to MitoPBN reacting with superoxide or the hydroxyl radical or by acting as a chain-breaking antioxidant. MitoPBN did react with carbon-centered radicals and also prevented lipid peroxidation by the carbon-centered radical generator 2,2'-azobis(2-methyl propionamidine) dihydrochloride (AAPH). Furthermore, AAPH activated UCPs, and this was blocked by MitoPBN. These data suggest that superoxide and lipid peroxidation products share a common pathway for the activation of UCPs. Superoxide releases iron from iron-sulfur center proteins, which then generates carbon-centered radicals that initiate lipid peroxidation, yielding breakdown products that activate UCPs.

  1. Mitochondria-targeting for improved photodynamic therapy

    NASA Astrophysics Data System (ADS)

    Ngen, Ethel J.

    Photodynamic therapy (PDT) is an emerging cancer therapeutic modality, with great potential to selectively treat surface cancers, thus minimizing systemic side effects. In this dissertation, two approaches to deliver photosensitizers to mitochondria were investigated: 1) Reducing photosensitizer sizes to improve endocytosis and lysosomal localization. Upon irradiation the photosensitizers would then produce singlet oxygen which could rupture the lysosomal membrane releasing the lysosomally trapped photosensitizers to the cytosol, from where they could relocalize to mitochondria by passive diffusion (photochemical internalization). 2) Using delocalized lipophilic cationic dyes (DLCs) to exploit membrane potential differences between the cytoplasm and mitochondria in delivering photosensitizers to mitochondria. To investigate the effects of steric hindrance on mitochondrial localization and photodynamic response, a series of eight thiaporphyrins were studied. Two new thiaporphyrin analogues 6 and 8 with reduced steric hindrance at the 10- and 15- meso positions were studied in comparison to 5,20-diphenyl-10,15-bis[4 (carboxymethyleneoxy)-phenyl]-21,23-dithiaporphyrin 1, previously validated as a potential second generation photosensitizer. Although 6 showed an extraordinarily high uptake (7.6 times higher than 1), it was less potent than 1 (IC 50 = 0.18 muM versus 0.13 muM) even though they both showed similar sub-cellular localization patterns. This low potency was attributed to its high aggregation tendency in aqueous media (4 times higher than 1), which might have affected its ability to generate singlet oxygen in vitro . 8 on the other hand showed an even lower potency than 6 (2.28 vs 0.18 muM). However this was attributed to its low cellular uptake (20 times less than 6) and inefficient generation of singlet oxygen. Overall, although the structural modifications did improve the cellular uptake of 6, 6 was still less potent than the lead photosensitizers 1. Thus, other strategies to target mitochondria for improved photodynamic activity were investigated. In a continuing project, we evaluated the ability of delocalized lipophilic cationic dyes to deliver photosensitizers to mitochondria by exploiting the membrane potential difference between the cytoplasm and mitochondria. Two conjugates: a porphyrin--rhodamine B conjugate (TPP--Rh) and a porphyrin-acridine orange conjugate (TPP--AO), each possessing a single delocalized lipophilic cation, were designed and synthesized. The conjugates were synthesized by conjugating a monohydroxy porphyrin (TPP-OH) to rhodamine B (Rh B) and acridine orange base (AO), respectively, via saturated hydrocarbon linkers. To evaluate the efficiency of the conjugates as photosensitizers, their photophysical properties and in vitro photodynamic activities were studied in comparison to those of TPP-OH, the parent porphyrin photosensitizer. Although fluorescence energy transfer (FRET) was observed in the conjugates, they were capable of generating singlet oxygen at rates comparable to TPP-OH. In a final project, we evaluated the photophysical potential of TPP-Rh to act as a two-photon photosensitizer for PDT. Two-photon PDT is a rational approach used to improve light penetration through the skin. Rhodamine B is an effective two-photon chromophore and could significantly improve the two-photon absorption of the porphyrin photosensitizer in the TPP-Rh dyad system following energy transfer. Thus the porphyrin--rhodamine B dyad (TPP--Rh), previously demonstrated to preferentially accumulate in the mitochondria, was photophysically evaluated as a potential two-photon photosensitizer. To evaluate the efficiency of TPP-Rh as a two-photon photosensitizer, its two-photon photophysical properties were compared with those of its individual components (Rh B and TPP-OH). This included: the two-photon cross sections (sigma 2), RET kinetics and dynamics and rates of singlet oxygen generation. A FRET efficiency of ~99 % was observed from the Rh moiety (donor) to the TPP moiety (acceptor) of the system. This significantly enhanced the sigma 2 of TPP-Rh by ˜ 100 % (20 GM) compared to the parent TPP-OH. Furthermore, TPP-Rh produced singlet oxygen at a significantly faster rate than TPP-OH upon two-photon excitation. Thus, this indicates that conjugating photosensitizers to Rh B via short saturated hydrocarbon linkers could provide deeper tissue penetration, in addition to preferential mitochondrial accumulation for improved photodynamic response. (Abstract shortened by UMI.)

  2. Kinetics and mechanism of hydroxyl radical and OH-adduct radical reactions with nitroxides and with their hydroxylamines.

    PubMed

    Samuni, Amram; Goldstein, Sara; Russo, Angelo; Mitchell, James B; Krishna, Murali C; Neta, Pedatsur

    2002-07-24

    Stable nitroxide radicals are potent antioxidants and are among the most effective non-thiol radioprotectants, although they react with hydroxyl radicals more slowly than typical phenolic antioxidants or thiols. Surprisingly, the reduced forms of cyclic nitroxides, cyclic hydroxylamines, are better reductants yet have no radioprotective activity. To clarify the reason for this difference, we studied the kinetics and mechanisms of the reactions of nitroxides and their hydroxylamines with (*)OH radicals and with OH-adducts by using pulse radiolysis, fluorimetric determination of phenolic radiation products, and electron paramagnetic resonance spectrometric determination of nitroxide concentrations following radiolysis. Competition kinetics with phenylalanine as a reference compound in pulse radiolysis experiments yielded rate constants of (4.5 +/- 0.4) x 10(9) M(-1) s(-1) for the reaction of (*)OH radical with 2,2,6,6-tetramethylpiperidine-N-oxyl (TPO), 4-hydroxy-TPO (4-OH-TPO), and 4-oxo-TPO (4-O-TPO), (3.0 +/- 0.3) x 10(9) M(-1) s(-1) for deuterated 4-O-TPO, and (1.0 +/- 0.1) x 10(9) M(-1) s(-1) for the hydroxylamine 4-OH-TPO-H. The kinetic isotope effect suggests the occurrence of both (*)OH addition to the aminoxyl moiety of 4-O-TPO and H-atom abstraction from the 2- or 6-methyl groups or from the 3- and 5-methylene positions. This conclusion was further supported by final product analysis, which demonstrated that (*)OH partially oxidizes 4-O-TPO to the corresponding oxoammonium cation. The rate constants for the reactions of the nitroxides with the OH-adducts of phenylalanine and terephthalate have been determined to be near 4 x 10(6) M(-1) s(-1), whereas the hydroxylamine reacted at least 50 times slower, if at all. These findings indicate that the reactivity toward (*)OH does not explain the differences between the radioprotective activities of nitroxides and hydroxylamines. Instead, the radioprotective activity of nitroxides, but not of hydroxylamines, can be

  3. The effects of derivatives of the nitroxide tempol on UVA-mediated in vitro lipid and protein oxidation.

    PubMed

    Damiani, Elisabetta; Castagna, Riccardo; Greci, Lucedio

    2002-07-01

    Derivatives of tetramethylpiperidines are extensively employed in polymers to prevent photooxidation, and their stabilizing effect is attributed to the activity of the nitroxide radical derived from the parent amine. In this study, we examined the photoprotective effect of a commercial polymer photostabilizer, HALS-1, its corresponding nitroxide, bis(2,2,6,6-tetramethyl-piperidine-1-oxyl-4-yl)sebacate (TINO), and two derivatives of the piperidine nitroxide TEMPOL, 2,2,6,6-tetramethyl-piperidin-4-acetyloxy-1-oxyl (TEMP2) and 2,2,6,6-tetramethyl-piperidin-4-octanoyloxy-1-oxyl (TEMP8) synthesized by us, in liposomes exposed to ultraviolet A (UVA) radiation. For comparison, the UVA-absorber, 4-tert-butyl-4'-methoxydibenzoylmethane (Parsol 1789) used in many suncream formulations, was also included. The nitroxide TINO resulted extremely efficient at inhibiting aldehydic breakdown products deriving from 30 min exposure of liposomes to UVA and the protection was dose-dependent (10-100 microM). The corresponding amine HALS-1 was the least efficient while protection increased in the order: TEMP2 < Parsol 1789 < TEMP 8. HALS-1, TINO, and the two TEMPOL derivatives were also tested in a simple protein system consisting of bovine serum albumin (BSA) exposed to UVA. In this case, these compounds did not inhibit nor enhance UVA-mediated protein carbonyl formation in BSA. The differences in protection between the compounds are discussed in relation to their chemical reactivity, UVA-absorbing capacities, and their molecular structure. Overall, the results obtained envisage the potential use of nitroxide compounds as topical antioxidants.

  4. Imaging of Nitroxides at 250 MHz using Rapid-Scan Electron Paramagnetic Resonance

    PubMed Central

    Biller, Joshua R.; Tseitlin, Mark; Quine, Richard W.; Rinard, George A.; Weismiller, Hilary A.; Elajaili, Hanan; Rosen, Gerald M.; Kao, Joseph P. Y.; Eaton, Sandra S.; Eaton, Gareth R.

    2014-01-01

    Projections for 2D spectral-spatial images were obtained by continuous wave and rapid-scan electron paramagnetic resonance using a bimodal cross-loop resonator at 251 MHz. The phantom consisted of three 4 mm tubes containing different 15N,2H-substituted nitroxides. Rapid-scan and continuous wave images were obtained with 5 min total acquisition times. For comparison, images also were obtained with 29 s acquisition time for rapid scan and 15 min for continuous wave. Relative to continuous wave projections obtained for the same data acquisition time, rapid-scan projections had significantly less low-frequency noise and substantially higher signal-to-noise at higher gradients. Because of the improved image quality for the same data acquisition time, linewidths could be determined more accurately from the rapid-scan images than from the continuous wave images. PMID:24650729

  5. Imaging of nitroxides at 250MHz using rapid-scan electron paramagnetic resonance.

    PubMed

    Biller, Joshua R; Tseitlin, Mark; Quine, Richard W; Rinard, George A; Weismiller, Hilary A; Elajaili, Hanan; Rosen, Gerald M; Kao, Joseph P Y; Eaton, Sandra S; Eaton, Gareth R

    2014-05-01

    Projections for 2D spectral-spatial images were obtained by continuous wave and rapid-scan electron paramagnetic resonance using a bimodal cross-loop resonator at 251MHz. The phantom consisted of three 4mm tubes containing different (15)N,(2)H-substituted nitroxides. Rapid-scan and continuous wave images were obtained with 5min total acquisition times. For comparison, images also were obtained with 29s acquisition time for rapid scan and 15min for continuous wave. Relative to continuous wave projections obtained for the same data acquisition time, rapid-scan projections had significantly less low-frequency noise and substantially higher signal-to-noise at higher gradients. Because of the improved image quality for the same data acquisition time, linewidths could be determined more accurately from the rapid-scan images than from the continuous wave images.

  6. Topology of exchange interactions in copper-nitroxide based molecular magnets studied by EPR

    NASA Astrophysics Data System (ADS)

    Drozdyuk, Irina Yu.; Maryunina, Ksenia Yu.; Sagdeev, Renad Z.; Ovcharenko, Victor I.; Bagryanskaya, Elena G.; Fedin, Matvey V.

    2013-10-01

    Switchable copper-nitroxide based molecular magnets Cu(hfac)2LR ('breathing crystals') have one-dimensional (1D) polymer-chain structure and 1D magnetic motif; however, the directions of polymer and magnetic chains in the crystal do not coincide. In this work we report the detailed electron paramagnetic resonance (EPR) study of the topology of intercluster exchange interactions forming the magnetic chains in a series of breathing crystals. The obtained results provide unambiguous manifestation of 1D magnetic behaviour, which degree correlates with the magnitude of intercluster exchange interaction. Analysis of experimental angular-dependent EPR data allowed us to determine relative orientations of magnetic chains with respect to polymer chains. The obtained angle between them is ≈40-46° for three studied compounds. The comparison of EPR and x-ray diffraction data confirms that 1D magnetic chains are formed by interacting adjacent spin triads of neighbouring polymer chains.

  7. Structures and magnetic properties of transition metal complexes involving 2,2'-bipyridin-6-yl nitroxide

    NASA Astrophysics Data System (ADS)

    Ondo, Akihiro; Ishida, Takayuki

    2017-01-01

    New complexes doubly chelated with two paramagnetic ligands, [MII(6bpyNO)2](ClO4)2•xCH3OH [M = Mn (x = 0.53), Ni (x = 1); 6bpyNO = 2,2'-bipyridin-6-yl tert-butyl nitroxide] were prepared. The X-ray crystallographic analysis revealed the isomorphous [M(6bpyNO)2]2+ structure. The magnetic measurements clarified antiferromagnetic 2JMn-rad/kB = -112(2) K and ferromagnetic 2JNi-rad/kB = +384(4) K, based on a symmetrical model H = -2JM-rad(SM•Srad1+SM•Srad2). The coupling mechanism is explained in terms of the 3d5 and 3d8 spin configurations leading to the dπ-pπoverlap in the Mn complex and the dσ-pπ orthogonal arrangement in the Ni complex along the metal-radical bonds.

  8. Nitroxide-mediated radical ring-opening copolymerization: chain-end investigation and block copolymer synthesis.

    PubMed

    Delplace, Vianney; Harrisson, Simon; Tardy, Antoine; Gigmes, Didier; Guillaneuf, Yohann; Nicolas, Julien

    2014-02-01

    Well-defined, degradable copolymers are successfully prepared by nitroxide-mediated radical ring opening polymerization (NMrROP) of oligo(ethylene glycol) methyl ether methacrylate (OEGMA) or methyl methacrylate (MMA), a small amount of acrylonitrile (AN) and cyclic ketene acetals (CKAs) of different structures. Phosphorous nuclear magnetic resonance allows in-depth chain-end characterization and gives crucial insights into the nature of the copoly-mer terminal sequences and the living chain fractions. By using a small library of P(OEGMA-co-AN-co-CKA) and P(MMA-co-AN-co-CKA) as macroinitiators, chain extensions with styrene are performed to furnish (amphiphilic) block copolymers comprising a degradable segment.

  9. Reactions of nitroxide radicals in aqueous solutions exposed to non-thermal plasma: limitations of spin trapping of the plasma induced species

    NASA Astrophysics Data System (ADS)

    Gorbanev, Yury; Stehling, Nicola; O'Connell, Deborah; Chechik, Victor

    2016-10-01

    Low temperature (‘cold’) atmospheric pressure plasmas have gained much attention in recent years due to their biomedical effects achieved through the interactions of plasma-induced species with the biological substrate. Monitoring of the radical species in an aqueous biological milieu is usually performed via electron paramagnetic resonance (EPR) spectroscopy using various nitrone spin traps, which form persistent radical adducts with the short-lived radicals. However, the stability of these nitroxide radical adducts in the plasma-specific environment is not well known. In this work, chemical transformations of nitroxide radicals in aqueous solutions using a model nitroxide 4-oxo-TEMPO were studied using EPR and LC-MS. The kinetics of the nitroxide decay when the solution was exposed to plasma were assessed, and the reactive pathways proposed. The use of different scavengers enabled identification of the types of reactive species which cause the decay, indicating the predominant nitroxide group reduction in oxygen-free plasmas. The 2H adduct of the PBN spin trap (PBN-D) was shown to decay similarly to the model molecule 4-oxo-TEMPO. The decay of the spin adducts in plasma-treated solutions must be considered to avoid rendering the spin trapping results unreliable. In particular, the selectivity of the decay indicated the limitations of the PTIO/PTI nitroxide system in the detection of nitric oxide.

  10. High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: Towards characterization of heterogeneity of microenvironments in spin-labeled systems

    NASA Astrophysics Data System (ADS)

    Nalepa, Anna; Möbius, Klaus; Lubitz, Wolfgang; Savitsky, Anton

    2014-05-01

    The combination of high-field EPR with site-directed spin-labeling (SDSL) techniques employing nitroxide radicals has turned out to be particularly powerful in probing the polarity and proticity characteristics of protein/matrix systems. This information is concluded from the principal components of the nitroxide Zeeman (g), nitrogen hyperfine (A) and quadrupole (P) tensors of the spin labels attached to specific sites. Recent multi-frequency high-field EPR studies underlined the complexity of the problem to treat the nitroxide microenvironment in proteins adequately due to inherent heterogeneities which result in several principal x-components of the nitroxide g-tensor. Concomitant, but distinctly different nitrogen hyperfine components could, however, not be determined from high-field cw EPR experiments owing to the large intrinsic EPR linewidth in fully protonated guest/host systems. It is shown in this work that, using the W-band (95 GHz) ELDOR- (electron-electron double resonance) detected NMR (EDNMR) method, different principal nitrogen hyperfine, Azz, and quadrupole, Pzz, tensor values of a nitroxide radical in glassy 2-propanol matrix can be measured with high accuracy. They belong to nitroxides with different hydrogen-bond situations. The satisfactory resolution and superior sensitivity of EDNMR as compared to the standard ENDOR (electron-nuclear double resonance) method are demonstrated.

  11. High-field ELDOR-detected NMR study of a nitroxide radical in disordered solids: towards characterization of heterogeneity of microenvironments in spin-labeled systems.

    PubMed

    Nalepa, Anna; Möbius, Klaus; Lubitz, Wolfgang; Savitsky, Anton

    2014-05-01

    The combination of high-field EPR with site-directed spin-labeling (SDSL) techniques employing nitroxide radicals has turned out to be particularly powerful in probing the polarity and proticity characteristics of protein/matrix systems. This information is concluded from the principal components of the nitroxide Zeeman (g), nitrogen hyperfine (A) and quadrupole (P) tensors of the spin labels attached to specific sites. Recent multi-frequency high-field EPR studies underlined the complexity of the problem to treat the nitroxide microenvironment in proteins adequately due to inherent heterogeneities which result in several principal x-components of the nitroxide g-tensor. Concomitant, but distinctly different nitrogen hyperfine components could, however, not be determined from high-field cw EPR experiments owing to the large intrinsic EPR linewidth in fully protonated guest/host systems. It is shown in this work that, using the W-band (95GHz) ELDOR- (electron-electron double resonance) detected NMR (EDNMR) method, different principal nitrogen hyperfine, Azz, and quadrupole, Pzz, tensor values of a nitroxide radical in glassy 2-propanol matrix can be measured with high accuracy. They belong to nitroxides with different hydrogen-bond situations. The satisfactory resolution and superior sensitivity of EDNMR as compared to the standard ENDOR (electron-nuclear double resonance) method are demonstrated.

  12. Effects of mitochondria-targeted plastoquinone derivative antioxidant (SkQ1) on demography of free-breeding Campbell dwarf hamsters (Phodopus campbelli) kept in outdoor conditions. reproduction and lifespan: explanation in the framework of ultimate loads.

    PubMed

    Rogovin, K A; Khrushcheva, A M; Shekarova, O N; Ushakova, M V; Manskikh, V N; Sokolova, O V; Vasilieva, N Yu

    2014-10-01

    We studied demographic effects of the mitochondria-targeted antioxidant SkQ1 on free-breeding Campbell dwarf hamsters (Phodopus campbelli, Thomas, 1905, Rodentia, Cricetidae) in an outdoor vivarium with seasonally varying day length and temperatures. The animals were kept in pairs from their young age. We removed litters from parental cages at their age of 25 days. Experimental hamsters received daily 50 nmol/kg SkQ1 with water by oral dosing, whereas control animals received water. SkQ1 had no effect on the lifespan of either males or females in reproductive pairs. Mortality among females was higher than among males irrespective of SkQ1 treatment, this being related to higher costs of reproduction in females. However, SkQ1 accelerated breeding in pairs in the first half of the reproductive period of a year. Although there were no statistical differences in body mass of males and females between experimental and control animals during most of their life, SkQ1-receiving males had higher body mass at the end of their life. The opposite tendency was characteristic for old females. One-year-old males and females of the experimental and control groups showed no difference in intensity of immune response to sheep red blood cells. The dermal hypersensitivity response to phytohemagglutinin (test for T-cell immunity) was significantly higher in SkQ1-treated 1- and 1.5-year-old males. This was not true for females. There was a tendency toward increased density of the neutrophil population in blood in 1-year-old SkQ1-treated males. However, experimental males showed no difference from control males in the activity of the "peroxidase-endogenous hydrogen peroxide system" of neutrophils. The background level of stress estimated by the concentration of cortisol in blood serum was significantly lower in the SkQ1-treated males during autumn adaptive adjustment of the organism. A similar trend was also observed during the January frosts, when the background level of stress was

  13. Controlled Redox Chemistry at Cerium within a Tripodal Nitroxide Ligand Framework

    SciTech Connect

    Bogart, Justin A.; Lippincott, Connor A.; Carroll, Patrick J.; Booth, Corwin H.; Schelter, Eric J.

    2015-10-27

    Ligand reorganization has been shown to have a profound effect on the outcome of cerium redox chemistry. Through the use of a tethered, tripodal, trianionic nitroxide ligand, [((2-tBuNOH)C 6 H 4 CH 2 ) 3 N] 3- (TriNO x 3- ), controlled redox chemistry at cerium was accomplished, and typically reactive complexes of tetravalent cerium were isolated. These included rare cationic complexes [Ce(TriNO x )thf][BAr F 4 ], in which Ar F =3,5-(CF 3 ) 2 -C 6 H 3 , and [Ce(TriNO x )py][OTf] . A rare complete Ce-halide series, Ce(TriNO x )X, in which X=F - , Cl - , Br - , I - , was also synthesized. We explored the solution chemistry of these complexes through detailed solution-phase electrochemistry and 1 H NMR experiments and showed a unique shift in the ratio of species with inner- and outer-sphere anions with size of the anionic X - group. DFT calculations on the series of calculations corroborated the experimental findings. Also, the use of a bulky and strongly donating tethered tripodal nitroxide ligand allowed the controlled redox chemistry at cerium. As a result, rare examples of cationic Ce IV complexes were synthesized and fully characterized. The full Ce-halide series supported by the tripodal ligand framework is also reported (see scheme).

  14. Translational diffusion in paramagnetic liquids by 1H NMR relaxometry: nitroxide radicals in solution.

    PubMed

    Kruk, D; Korpała, A; Kubica, A; Meier, R; Rössler, E A; Moscicki, J

    2013-01-14

    For nitroxide radicals in solution one can identify three frequency regimes in which (1)H spin-lattice relaxation rate of solvent molecules depend linearly on square root of the (1)H resonance frequency. Combining a recently developed theory of nuclear (proton) spin-lattice relaxation in solutions of nitroxide radicals [D. Kruk et al., J. Chem. Phys. 137, 044512 (2012)] with properties of the spectral density function associated with translational dynamics, relationships between the corresponding linear changes of the relaxation rate (for (14)N spin probes) and relative translational diffusion coefficient of the solvent and solute molecules have been derived (in analogy to (15)N spin probes [E. Belorizky et al., J. Phys. Chem. A 102, 3674 (1998)]). This method allows a simple and straightforward determination of diffusion coefficients in spin-labeled systems, by means of (1)H nuclear magnetic resonance (NMR) relaxometry. The approach has thoroughly been tested by applying to a large set of experimental data-(1)H spin-lattice relaxation dispersion results for solutions of different viscosity (decalin, glycerol, propylene glycol) of (14)N and (15)N spin probes. The experiments have been performed versus temperature (to cover a broad range of translational diffusion coefficients) using field cycling spectrometer which covers three decades in (1)H resonance frequency, 10 kHz-20 MHz. The limitations of NMR relaxometry caused by the time scale of the translational dynamics as well as electron spin relaxation have been discussed. It has been shown that for spin-labeled systems NMR relaxometry gives access to considerably faster diffusion processes than for diamagnetic systems.

  15. Translational diffusion in paramagnetic liquids by 1H NMR relaxometry: Nitroxide radicals in solution

    NASA Astrophysics Data System (ADS)

    Kruk, D.; Korpała, A.; Kubica, A.; Meier, R.; Rössler, E. A.; Moscicki, J.

    2013-01-01

    For nitroxide radicals in solution one can identify three frequency regimes in which 1H spin-lattice relaxation rate of solvent molecules depend linearly on square root of the 1H resonance frequency. Combining a recently developed theory of nuclear (proton) spin-lattice relaxation in solutions of nitroxide radicals [D. Kruk et al., J. Chem. Phys. 137, 044512 (2012)], 10.1063/1.4736854 with properties of the spectral density function associated with translational dynamics, relationships between the corresponding linear changes of the relaxation rate (for 14N spin probes) and relative translational diffusion coefficient of the solvent and solute molecules have been derived (in analogy to 15N spin probes [E. Belorizky et al., J. Phys. Chem. A 102, 3674 (1998)], 10.1021/jp980397h). This method allows a simple and straightforward determination of diffusion coefficients in spin-labeled systems, by means of 1H nuclear magnetic resonance (NMR) relaxometry. The approach has thoroughly been tested by applying to a large set of experimental data—1H spin-lattice relaxation dispersion results for solutions of different viscosity (decalin, glycerol, propylene glycol) of 14N and 15N spin probes. The experiments have been performed versus temperature (to cover a broad range of translational diffusion coefficients) using field cycling spectrometer which covers three decades in 1H resonance frequency, 10 kHz-20 MHz. The limitations of NMR relaxometry caused by the time scale of the translational dynamics as well as electron spin relaxation have been discussed. It has been shown that for spin-labeled systems NMR relaxometry gives access to considerably faster diffusion processes than for diamagnetic systems.

  16. Nitroxides as anti-biofilm compounds for the treatment of Pseudomonas aeruginosa and mixed-culture biofilms.

    PubMed

    Alexander, Stefanie-Ann; Kyi, Caroline; Schiesser, Carl H

    2015-04-28

    A series of 23 nitroxides () was tested for biofilm modulatory activity using a crystal violet staining technique. 3-(Dodecane-1-thiyl)-4-(hydroxymethyl)-2,2,5,5-tetramethyl-1-pyrrolinoxyl () was found to significantly suppress biofilm formation and elicit dispersal events in both Pseudomonas aeruginosa and mixed-culture biofilms. Twitching and swarming motilities were enhanced by nitroxide , leaving the planktonic-specific swimming motility unaffected and suggesting that the mechanism of -mediated biofilm modulation is linked to the hyperactivation of surface-associated cell motilities. Preliminary structure-activity relationship studies identify the dodecanethiyl chain, hydroxymethyl substituent and the free radical moiety to be structural features pertinent to the anti-biofilm activity of .

  17. Determination of the 14N quadrupole coupling constant of nitroxide spin probes by W-band ELDOR-detected NMR.

    PubMed

    Florent, Marc; Kaminker, Ilia; Nagarajan, Vijayasarathi; Goldfarb, Daniella

    2011-06-01

    Nitroxide spin probe electron paramagnetic resonance (EPR) has proven to be a very successful method to probe local polarity and solvent hydrogen bonding properties at the molecular level. The g(xx) and the (14)N hyperfine A(zz) principal values are the EPR parameters of the nitroxide spin probe that are sensitive to these properties and are therefore monitored experimentally. Recently, the (14)N quadrupole interaction of nitroxides has been shown to be also highly sensitive to polarity and H-bonding (A. Savitsky et al., J. Phys. Chem. B 112 (2008) 9079). High-field electron spin echo envelope modulation (ESEEM) was used successfully to determine the P(xx) and P(yy) principal components of the (14)N quadrupole tensor. The P(zz) value was calculated from the traceless character of the quadrupole tensor. We introduce here high-field (W-band, 95 GHz, 3.5 T) electron-electron double resonance (ELDOR)-detected NMR as a method to obtain the (14)N P(zz) value directly, together with A(zz). This is complemented by W-band hyperfine sublevel correlation (HYSCORE) measurements carried out along the g(xx) direction to determine the principal P(xx) and P(yy) components. Through measurements of TEMPOL dissolved in solvents of different polarities, we show that A(zz) increases, while |P(zz)| decreases with polarity, as predicted by Savitsky et al.

  18. Determination of the 14N quadrupole coupling constant of nitroxide spin probes by W-band ELDOR-detected NMR

    NASA Astrophysics Data System (ADS)

    Florent, Marc; Kaminker, Ilia; Nagarajan, Vijayasarathi; Goldfarb, Daniella

    2011-06-01

    Nitroxide spin probe electron paramagnetic resonance (EPR) has proven to be a very successful method to probe local polarity and solvent hydrogen bonding properties at the molecular level. The g xx and the 14N hyperfine A zz principal values are the EPR parameters of the nitroxide spin probe that are sensitive to these properties and are therefore monitored experimentally. Recently, the 14N quadrupole interaction of nitroxides has been shown to be also highly sensitive to polarity and H-bonding (A. Savitsky et al., J. Phys. Chem. B 112 (2008) 9079). High-field electron spin echo envelope modulation (ESEEM) was used successfully to determine the P xx and P yy principal components of the 14N quadrupole tensor. The P zz value was calculated from the traceless character of the quadrupole tensor. We introduce here high-field (W-band, 95 GHz, 3.5 T) electron-electron double resonance (ELDOR)-detected NMR as a method to obtain the 14N P zz value directly, together with A zz. This is complemented by W-band hyperfine sublevel correlation (HYSCORE) measurements carried out along the g xx direction to determine the principal P xx and P yy components. Through measurements of TEMPOL dissolved in solvents of different polarities, we show that A zz increases, while | P zz| decreases with polarity, as predicted by Savitsky et al.

  19. A Nitroxide-Tagged Platinum(II) Complex Enables the Identification of DNA G-Quadruplex Binding Mode

    PubMed Central

    Zhou, Yi-Wei; Wang, Hanqiang; Cao, Qian; Shen, Yong; Ji, Liang-Nian; Mao, Zong-Wan; Qin, Peter Z.

    2016-01-01

    We reported a novel strategy for investigating small molecule binding to G-quadruplexes (GQs). A newly synthesized dinuclear platinum(II) complex (Pt2L) containing a nitroxide radical was shown to selectively bind a GQ-forming sequence derived from human telomere (hTel). Using the nitroxide moiety as a spin label, electron paramagnetic resonance (EPR) spectroscopy was carried out to investigate binding between Pt2L and hTel GQ. Measurements indicated that two molecules of Pt2L bind with one molecule of hTel GQ. The inter-spin distance measured between the two bound Pt2L, together with molecular docking analyses, revealed that Pt2L predominately binds to the neighboring narrow and wide grooves of the G-tetrads as hTel adopts the antiparallel conformation. The design and synthesis of nitroxide tagged GQ binders, and the use of spin-labeling/EPR to investigate their interactions with GQs, will aid the development of small molecules for manipulating GQs involved in crucial biological processes. PMID:26845489

  20. In vivo evaluation of different alterations of redox status by studying pharmacokinetics of nitroxides using magnetic resonance techniques

    PubMed Central

    Bačić, Goran; Pavićević, Aleksandra; Peyrot, Fabienne

    2015-01-01

    Free radicals, particularly reactive oxygen species (ROS), are involved in various pathologies, injuries related to radiation, ischemia-reperfusion or ageing. Unfortunately, it is virtually impossible to directly detect free radicals in vivo, but the redox status of the whole organism or particular organ can be studied in vivo by using magnetic resonance techniques (EPR and MRI) and paramagnetic stable free radicals – nitroxides. Here we review results obtained in vivo following the pharmacokinetics of nitroxides on experimental animals (and a few in humans) under various conditions. The focus was on conditions where the redox status has been altered by induced diseases or harmful agents, clearly demonstrating that various EPR/MRI/nitroxide combinations can reliably detect metabolically induced changes in the redox status of organs. These findings can improve our understanding of oxidative stress and provide a basis for studying the effectiveness of interventions aimed to modulate oxidative stress. Also, we anticipate that the in vivo EPR/MRI approach in studying the redox status can play a vital role in the clinical management of various pathologies in the years to come providing the development of adequate equipment and probes. PMID:26827126

  1. Synthesis of a novel adamantyl nitroxide derivative with potent anti-hepatoma activity in vitro and in vivo.

    PubMed

    Sun, Jin; Wang, Shan; Bu, Wei; Wei, Meng-Ying; Li, Wei-Wei; Yao, Min-Na; Ma, Zhong-Ying; Lu, Cheng-Tao; Li, Hui-Hui; Hu, Na-Ping; Zhang, En-Hu; Yang, Guo-Dong; Wen, Ai-Dong; Zhu, Xiao-He

    2016-01-01

    In this study, a novel adamantyl nitroxide derivative was synthesized and its antitumor activities in vitro and in vivo were investigated. The adamantyl nitroxide derivative 4 displayed a potent anticancer activity against all the tested human hepatoma cells, especially with IC50 of 68.1 μM in Bel-7404 cells, compared to the positive control 5-FU (IC50=607.7 μM). The significant inhibition of cell growth was also observed in xenograft mouse model, with low toxicity. Compound 4 suppressed the cell migration and invasion, induced the G2/M phase arrest. Further mechanistic studies revealed that compound 4 induced cell death, which was accompanied with damaging mitochondria, increasing the generation of intracellular reactive oxygen species, cleavages of caspase-9 and caspase-3, as well as activations of Bax and Bcl-2. These results confirmed that adamantyl nitroxide derivative exhibited selective antitumor activities via mitochondrial apoptosis pathway in Bel-7404 cells, and would be a potential anticancer agent for liver cancer.

  2. Synthesis of a novel adamantyl nitroxide derivative with potent anti-hepatoma activity in vitro and in vivo

    PubMed Central

    Sun, Jin; Wang, Shan; Bu, Wei; Wei, Meng-Ying; Li, Wei-Wei; Yao, Min-Na; Ma, Zhong-Ying; Lu, Cheng-Tao; Li, Hui-Hui; Hu, Na-Ping; Zhang, En-Hu; Yang, Guo-Dong; Wen, Ai-Dong; Zhu, Xiao-He

    2016-01-01

    In this study, a novel adamantyl nitroxide derivative was synthesized and its antitumor activities in vitro and in vivo were investigated. The adamantyl nitroxide derivative 4 displayed a potent anticancer activity against all the tested human hepatoma cells, especially with IC50 of 68.1 μM in Bel-7404 cells, compared to the positive control 5-FU (IC50=607.7 μM). The significant inhibition of cell growth was also observed in xenograft mouse model, with low toxicity. Compound 4 suppressed the cell migration and invasion, induced the G2/M phase arrest. Further mechanistic studies revealed that compound 4 induced cell death, which was accompanied with damaging mitochondria, increasing the generation of intracellular reactive oxygen species, cleavages of caspase-9 and caspase-3, as well as activations of Bax and Bcl-2. These results confirmed that adamantyl nitroxide derivative exhibited selective antitumor activities via mitochondrial apoptosis pathway in Bel-7404 cells, and would be a potential anticancer agent for liver cancer. PMID:27429843

  3. The role of charge-transfer interactions and delocalization in annelated nitronyl nitroxides

    NASA Astrophysics Data System (ADS)

    Dooley, Brynn Mary

    The design and synthesis of stable organic radicals with delocalized spin density distribution and low energy optical and redox processes is central to the development of magneto-conducting materials. Towards this end, a generalized synthetic methodology has been developed allowing for the synthesis of a series of annelated benzonitronyl nitroxide (BNN) radicals. The BNN radicals exhibited remarkably low reduction potentials (~0.0 V vs SCE) and a near-infrared absorption attributed to a HOMO--SOMO charge-transfer excitation. The annelated BNN radicals were found to be less stable than the closely related tetramethyl nitronyl nitroxide radicals, particularly in solution. A series of pi-delocalized and heteroaromatic radicals were synthesized to determine if the instability was due to the delocalization of electron density onto the carbon skeleton or the low reduction potential. DFT calculations with the EPR-II basis gave rise to calculated electronic structures that were consistent with EPR spectroscopy and suggested changes in spin density distribution are occurring with perturbation of the annelated ring. Cyclic voltammetry revealed the heteroaromatic and pi-delocalized radicals had reduction potentials lower than BNN, with some systems reducing at potentials of 0.2 V vs SCE, comparable to that of 7,7,8,8-tetracyanoquinodimethane. The distribution of spin throughout the molecular framework and the low reduction potential of the annelated nitronyl nitroxide radicals were both found to contribute to the lowered stability of the annelated nitronyl nitroxides relative to the far less redox active tetramethyl nitronyl nitroxides. The low reduction potential of the BNN radicals suggested that incorporation into acceptor--donor (A--D) systems would allow for investigation of the role of charge transfer interactions on the electronic structure and properties of neutral open-shell A--D radicals. Two D--A--D radicals were prepared using metal catalyzed coupling and furoxan

  4. Tuning the Spin Coupling Interactions in the Nitroxide-Based Bisphenol-Like Diradicals.

    PubMed

    Song, Meiyu; Song, Xinyu; Bu, Yuxiang

    2017-07-24

    The intramolecular spin coupling interactions of bisphenol-like trinary-bridged diradicals [nitroxide-(para/meta)phenylene-X-phenylene(para/meta)-nitroxide, X=C=CH2 , O, BH, NH and SO2 ] were explored with an emphasis on the tuning role of the X coupler at the (U)B3LYP/6-311++G(d,p) level. Our results indicate that all designed trinary-bridged diradicals featuring a V-type structure with a bending angle of 104-130° and torsional angles of two phenylene rings being 20-90° exhibit different diradical character and magnetism, depending on the structures and properties of the X bridges. More interestingly, although meta/para-phenylene supports a ferromagnetic (FM)/antiferromagnetic (AFM) coupling, their combinations by using X as a trinary bridge can mediate spin coupling, but the coupling magnitude strongly depends on X. In general, a para/para or meta/meta combination with X leads to an open-shell singlet ground state and thus AFM, but the meta/meta combination considerably decreases the spin coupling interaction. In contrast, a para/meta combination with X produces a triplet ground state and FM. Their combination with a single-electron conjugation end coupler (C=CH2 ) leads to an inverse coupling regularity. All results can be reasonably explained by the spin alternation rule, molecular structures, and properties. In particular, detailed spin coupling mechanisms are suggested to involve cooperative through-space and through-bond pathways with different levels of cooperativity. This is rationalized with the X-induced bending of the diradicals not only modifying the through-bond (extended π conjugation) pathway but also provididng a through-space (face-to-face vs. side-to-side π-π interaction) possibility for spin coupling, in conjunction with twisting of the phenylene rings. Different contributions of the through-space and through-bond couplings are quantitatively distinguished and depend on the structure and property of the X coupler. Clearly, this work reports

  5. The nitroxide Tempo inhibits hydroxyl radical production from the Fenton-like reaction of iron(II)-citrate with hydrogen peroxide.

    PubMed

    Shi, Fengqiang; Zhang, Peifeng; Mao, Yujia; Wang, Can; Zheng, Meiqing; Zhao, Zhongwei

    2017-01-29

    In vivo physiological ligand citrate can bind iron(II) ions to form the iron(II)-citrate complex. Inhibition of hydroxyl radical (OH) production from the Fenton-like reaction of iron(II)-citrate with H2O2 is biologically important, as this reaction may account for one of the mechanisms of the labile iron pool in vivo to induce oxidative stress and pathological conditions. Nitroxides have promising potentials as therapeutic antioxidants. However, there are controversial findings indicating that they not only act as antioxidants but also as pro-oxidants when engaged in Fenton reactions. Although the underlying mechanisms are proposed to be the inhibition or enhancement of the OH production by nitroxides, the proposed elucidations are only based on assessing biological damages and not demonstrated directly by measuring the OH production in the presence of nitroxides. In this study, therefore, we employed EPR and fluorescence spectroscopies to show direct evidence that nitroxide 2,2,6,6-tetramethyl-piperidine-1-oxyl (Tempo) inhibited OH production from the Fenton-like reaction of iron(II)-citrate with H2O2 by up to 90%. We also demonstrated spectrophotometrically, for the first time, that this inhibition was due to oxidation of the iron(II)-citrate by Tempo with a stoichiometry of Tempo:Iron(III)-citrate = 1.1:1.0. A scheme was proposed to illustrate the roles of nitroxides engaged in Fenton/Fenton-like reactions.

  6. Nitrogen nuclear spin flips in nitroxide spin probes of different sizes in glassy o-terphenyl: Possible relation with α- and β-relaxations

    NASA Astrophysics Data System (ADS)

    Isaev, N. P.; Dzuba, S. A.

    2011-09-01

    The pulsed electron-electron double resonance (ELDOR) technique was employed to study nitroxide spin probes of three different sizes dissolved in glassy o-terphenyl. A microwave pulse applied to the central hyperfine structure (hfs) component of the nitroxide electron paramagnetic resonance spectrum was followed by two echo-detecting pulses of different microwave frequency to probe the magnetization transfer (MT) to the low-field hfs component. The MT between hfs components is readily related to flips in the nitrogen nuclear spin, which in turn are induced by molecular motion. The MT on the time scale of tens of microseconds was observed over a wide temperature range, including temperatures near and well below the glass transition. For a bulky nitroxide, it was found that MT rates approach dielectric α (primary) relaxation frequencies reported for o-terphenyl in the literature. For small nitroxides, MT rates were found to match the frequencies of dielectric β (secondary) Johari-Goldstein relaxation. The most probable motional mechanism inducing the nitrogen nuclear spin flips is large-angle angular jumps, between some orientations of unequal occupation probabilities. The pulsed ELDOR of nitroxide spin probes may provide additional insight into the nature of Johari-Goldstein relaxation in glassy media and may serve as a tool for studying this relaxation in substances consisting of non-rigid molecules (such as branched polymers) and in heterogeneous and non-polar systems (such as a core of biological membranes).

  7. Rationalizing Overhauser DNP of nitroxide radicals in water through MD simulations.

    PubMed

    Sezer, Deniz

    2014-01-21

    The recently introduced methodology (Sezer, Phys. Chem. Chem. Phys., 2013, 15, 526) for calculating dynamic nuclear polarization (DNP) coupling factors through synergistic use of molecular dynamics (MD) simulations and the analytical model of diffusing hard spheres with spins at their centers (HSCS) is applied to several nitroxides in water. Computations with one TEMPONE and one TEMPOL in water agree with experiments at 0.35 T and 3.4 T, respectively. At 9.2 T the predicted coupling factors are larger by about 50% than the experimental numbers obtained with 1 M TEMPOL solution. MD simulations at this elevated concentration reveal nanoscopic TEMPOL clusters and qualitatively explain the lower experimental values. Comparing the dynamics from the MD simulations with those of the HSCS model, the assumption of centered spins is shown to be too limiting even for small molecules like TEMPOL and water. Using the available extension of the HSCS model to off-centered spins, the current procedure for analyzing hydration water dynamics from Overhauser DNP measurements on spin-labeled proteins is revisited.

  8. Mitochondrial dysfunction and cell death in neurodegenerative diseases through nitroxidative stress.

    PubMed

    Akbar, Mohammed; Essa, Musthafa Mohamed; Daradkeh, Ghazi; Abdelmegeed, Mohamed A; Choi, Youngshim; Mahmood, Lubna; Song, Byoung-Joon

    2016-04-15

    Mitochondria are important for providing cellular energy ATP through the oxidative phosphorylation pathway. They are also critical in regulating many cellular functions including the fatty acid oxidation, the metabolism of glutamate and urea, the anti-oxidant defense, and the apoptosis pathway. Mitochondria are an important source of reactive oxygen species leaked from the electron transport chain while they are susceptible to oxidative damage, leading to mitochondrial dysfunction and tissue injury. In fact, impaired mitochondrial function is commonly observed in many types of neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, Huntington's disease, alcoholic dementia, brain ischemia-reperfusion related injury, and others, although many of these neurological disorders have unique etiological factors. Mitochondrial dysfunction under many pathological conditions is likely to be promoted by increased nitroxidative stress, which can stimulate post-translational modifications (PTMs) of mitochondrial proteins and/or oxidative damage to mitochondrial DNA and lipids. Furthermore, recent studies have demonstrated that various antioxidants, including naturally occurring flavonoids and polyphenols as well as synthetic compounds, can block the formation of reactive oxygen and/or nitrogen species, and thus ultimately prevent the PTMs of many proteins with improved disease conditions. Therefore, the present review is aimed to describe the recent research developments in the molecular mechanisms for mitochondrial dysfunction and tissue injury in neurodegenerative diseases and discuss translational research opportunities.

  9. Stabilization of reactive nitroxides using invasomes to allow prolonged electron paramagnetic resonance measurements.

    PubMed

    Haag, S F; Taskoparan, B; Bittl, R; Teutloff, C; Wenzel, R; Fahr, A; Chen, M; Lademann, J; Schäfer-Korting, M; Meinke, M C

    2011-01-01

    The detection of the antioxidative capacity of the skin is of great practical relevance since free radicals are involved in many skin damaging processes, including aging and inflammation. The nitroxide TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxyl) in combination with electron paramagnetic resonance spectroscopy was found suitable for measuring the antioxidative capacity since its reaction with reducing agents is considerably fast. Yet, in order to achieve longer measurement times, e.g. in inflammatory skin diseases, the stabilizing effect of an invasome (ultraflexible vesicle/liposome) suspension with TEMPO was investigated ex vivo on porcine skin and in vivo on human skin. Invasomes increased the measurement time ex vivo 2-fold and the reduction was significantly slowed down in vivo, which is due to membrane-associated and therefore protected TEMPO. Furthermore, TEMPO accumulation in the membrane phase as well as the decreasing polarity of the ultimate surroundings of TEMPO during skin penetration explains the stabilizing effect. Thus, an invasome suspension with TEMPO exhibits stabilizing effects ex vivo and in vivo.

  10. A Mitochondrial-Targeted Nitroxide Is a Potent Inhibitor of Ferroptosis.

    PubMed

    Krainz, Tanja; Gaschler, Michael M; Lim, Chaemin; Sacher, Joshua R; Stockwell, Brent R; Wipf, Peter

    2016-09-28

    Discovering compounds and mechanisms for inhibiting ferroptosis, a form of regulated, nonapoptotic cell death, has been of great interest in recent years. In this study, we demonstrate the ability of XJB-5-131, JP4-039, and other nitroxide-based lipid peroxidation mitigators to prevent ferroptotic cell death in HT-1080, BJeLR, and panc-1 cells. Several analogues of the reactive oxygen species (ROS) scavengers XJB-5-131 and JP4-039 were synthesized to probe structure-activity relationships and the influence of subcellular localization on the potency of these novel ferroptosis suppressors. Their biological activity correlated well over several orders of magnitude with their structure, relative lipophilicity, and respective enrichment in mitochondria, revealing a critical role of intramitochondrial lipid peroxidation in ferroptosis. These results also suggest that preventing mitochondrial lipid oxidation might offer a viable therapeutic opportunity in ischemia/reperfusion-induced tissue injury, acute kidney injury, and other pathologies that involve ferroptotic cell death pathways.

  11. A Mitochondrial-Targeted Nitroxide Is a Potent Inhibitor of Ferroptosis

    PubMed Central

    2016-01-01

    Discovering compounds and mechanisms for inhibiting ferroptosis, a form of regulated, nonapoptotic cell death, has been of great interest in recent years. In this study, we demonstrate the ability of XJB-5-131, JP4-039, and other nitroxide-based lipid peroxidation mitigators to prevent ferroptotic cell death in HT-1080, BJeLR, and panc-1 cells. Several analogues of the reactive oxygen species (ROS) scavengers XJB-5-131 and JP4-039 were synthesized to probe structure–activity relationships and the influence of subcellular localization on the potency of these novel ferroptosis suppressors. Their biological activity correlated well over several orders of magnitude with their structure, relative lipophilicity, and respective enrichment in mitochondria, revealing a critical role of intramitochondrial lipid peroxidation in ferroptosis. These results also suggest that preventing mitochondrial lipid oxidation might offer a viable therapeutic opportunity in ischemia/reperfusion-induced tissue injury, acute kidney injury, and other pathologies that involve ferroptotic cell death pathways. PMID:27725964

  12. Analysis of nitroxide spin label motion in a protein protein complex using multiple frequency EPR spectroscopy

    NASA Astrophysics Data System (ADS)

    White, G. F.; Ottignon, L.; Georgiou, T.; Kleanthous, C.; Moore, G. R.; Thomson, A. J.; Oganesyan, V. S.

    2007-04-01

    X- and W-band EPR spectra, at room and low temperatures, are reported for nitroxide spin labels attached to cysteine residues selectively introduced into two proteins, the DNase domain of colicin-E9 and its immunity protein, Im9. The dynamics of each site of attachment on the individual proteins and in the tight DNase-Im9 complex have been analysed by computer simulations of the spectra using a model of Brownian dynamics trajectories for the spin label and protein. Ordering potentials have been introduced to describe mobility of labels restricted by the protein domain. Label mobility varies with position from completely immobilised, to motionally restricted and to freely rotating. Bi-modal dynamics of the spin label have been observed for several sites. We show that W-band spectra are particularly useful for detection of anisotropy of spin label motion. On complex formation significant changes are observed in the dynamics of labels at the binding interface region. This work reveals multi-frequency EPR as a sensitive and valuable tool for detecting conformational changes in protein structure and dynamics especially in protein-protein complexes.

  13. Metal-free and MRI visible theranostic lyotropic liquid crystal nitroxide-based nanoparticles.

    PubMed

    Muir, Benjamin W; Acharya, Durga P; Kennedy, Danielle F; Mulet, Xavier; Evans, Richard A; Pereira, Suzanne M; Wark, Kim L; Boyd, Ben J; Nguyen, Tri-Hung; Hinton, Tracey M; Waddington, Lynne J; Kirby, Nigel; Wright, David K; Wang, Hong X; Egan, Gary F; Moffat, Bradford A

    2012-03-01

    The development of improved, low toxicity, clinically viable nanomaterials that provide MRI contrast have tremendous potential to form the basis of translatable theranostic agents. Herein we describe a class of MRI visible materials based on lyotropic liquid crystal nanoparticles loaded with a paramagnetic nitroxide lipid. These readily synthesized nanoparticles achieved enhanced proton-relaxivities on the order of clinically used gadolinium complexes such as Omniscan™ without the use of heavy metal coordination complexes. Their low toxicity, high water solubility and colloidal stability in buffer resulted in them being well tolerated in vitro and in vivo. The nanoparticles were initially screened in vitro for cytotoxicity and subsequently a defined concentration range was tested in rats to determine the maximum tolerated dose. Pharmacokinetic profiles of the candidate nanoparticles were established in vivo on IV administration to rats. The lyotropic liquid crystal nanoparticles were proven to be effective liver MRI contrast agents. We have demonstrated the effective in vivo performance of a T1 enhancing, biocompatible, colloidally stable, amphiphilic MRI contrast agent that does not contain a metal. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Nitroxide TEMPO: a genotoxic and oxidative stress inducer in cultured cells.

    PubMed

    Guo, Xiaoqing; Mittelstaedt, Roberta A; Guo, Lei; Shaddock, Joseph G; Heflich, Robert H; Bigger, Anita H; Moore, Martha M; Mei, Nan

    2013-08-01

    2,2,6,6-Tetramethylpiperidine-1-oxyl (TEMPO) is a low molecular weight nitroxide and stable free radical. In this study, we investigated the cytotoxicity and genotoxicity of TEMPO in mammalian cells using the mouse lymphoma assay (MLA) and in vitro micronucleus assay. In the absence of metabolic activation (S9), 3mM TEMPO produced significant cytotoxicity and marginal mutagenicity in the MLA; in the presence of S9, treatment of mouse lymphoma cells with 1-2mM TEMPO resulted in dose-dependent decreases of the relative total growth and increases in mutant frequency. Treatment of TK6 human lymphoblastoid cells with 0.9-2.3mM TEMPO increased the frequency of both micronuclei (a marker for clastogenicity) and hypodiploid nuclei (a marker of aneugenicity) in a dose-dependent manner; greater responses were produced in the presence of S9. Within the dose range tested, TEMPO induced reactive oxygen species and decreased glutathione levels in mouse lymphoma cells. In addition, the majority of TEMPO-induced mutants had loss of heterozygosity at the Tk locus, with allele loss of ⩽34Mbp. These results indicate that TEMPO is mutagenic in the MLA and induces micronuclei and hypodiploid nuclei in TK6 cells. Oxidative stress may account for part of the genotoxicity induced by TEMPO in both cell lines.

  15. Syntheses, structure, magnetic and thermodynamics property of novel lanthanide complexes with nitronyl nitroxide radical

    NASA Astrophysics Data System (ADS)

    Song, Mei-Ying; Hou, Yi-Fang; Wen, Long-Mei; Wang, Shu-Ping; Yang, Shu-Tao; Zhang, Jian-Jun; Geng, Li-Na; Shi, Shi-Kao

    2016-03-01

    Four new nitronyl nitroxide radical-Ln(III) complexes, Ln(hfac)3(NITPhSCF3)2 (Ln(III) = Sm(1), Gd(2), Tb(3), Dy(4); NITPhSCF3 = 2-(4-trifluoromethylthiophenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl- 3-oxide; hfac = hexafluoroacetylacetonate), have been synthesized and characterized. They are isostructural, which show mononuclear tri-spin structures. The central Ln(III) ion is eight-coordinated by three hfac anions and two NITPhSCF3 molecules. Direct-current magnetic study shows that there exist ferromagnetic interactions between Gd(III) ion and radicals (NITPhSCF3) with JGd-Rad = 1.61 cm-1, and antiferromagnetic interactions between radicals with JRad-Rad = -2.83 cm-1 in complex 2. The magnetic analysis with the rough approximate model show that a ferromagnetic coupling exists between Tb(III) and radical in 3, while a antiferromagnetic coupling between Dy(III) and radical in 4. The thermodynamics properties of four complexes were studied with differential scanning calorimetry (DSC), such as heat capacity, thermodynamic functions (HT-H298.15K), (ST-S298.15K), and (GT-G298.15K).

  16. Effects of oxygen on EPR spectra of nitroxide spin-label probes of model membranes

    NASA Astrophysics Data System (ADS)

    Popp, Carol A.; Hyde, James S.

    The use of a methylpentene polymer, TPX, for construction of sample containers that allow easy equilibration of electron paramagnetic resonance samples with nitrogen is described. The effects of oxygen-dependent shortening of the electron spin relaxation times of nitroxide spin labels were studied in dispersions of dimyristoylphosphatidylcholine (DMPC) and dipalmitoylphosphatidylchohne (DPPC). First-harmonic, in-phase, absorption spectra of deoxygenated samples of 2-(14-carboxytetradecyl)-2-ethyl-4,4-dimethyl-3-oxazolidinyloxyl (16SASL) in DMPC display decreased linewidths and increased peak-to-peak heigths and resolution of 13C splittings. Continuous-wave (cw) saturation studies of 16SASL/DMPC and both lipid- and aqueous-phase components of 2,2,6,6-tetramethyl-piperidinooxyl (Tempo) partitioned into DPPC show that the rf field at which the signal intensity is maximized decreases when aerated samples are equilibrated with nitrogen. Second-harmonic, out-of-phase, absorption (saturation transfer) spectra of deoxygenated samples of 16SASUDMPC at -22°C and 2-(3-carboxypropyl)-4,4-dimethyl-2-tridecyl-3-oxazolidinyloxyl (5SASL) in DPPC at 35°C display increased signal intensity and lineshape changes. Electron-electron double resonance (ELDOR) spectra display much greater ELDOR reduction in signal intensity when a deoxygenated sample of 16SASL/DMPC is used. Our results indicate that the routine use of deoxygenated samples in biologically relevant studies using spin-label probes should be considered.

  17. 2D-ELDOR detection of magnetization transfer of nitroxides in disordered solid polymers

    NASA Astrophysics Data System (ADS)

    Maresch, G. G.; Weber, M.; Dubinskii, A. A.; Spiess, H. W.

    1992-05-01

    Two-dimensional electron—electron double resonance (2D-ELDOR) experiments on nitroxide spin labels in solid liquid-crystalline side-group polymers have been performed employing narrow-band microwave excitation pulses followed by a rapid magnetic field step during a mixing time and detection at the new selected point of the EPR spectrum. Information about magnetization transfer throughout the full EPR spectrum is obtained by sweeping both pumping and detecting fields. In the two-dimensional representation of experimental ELDOR data, the different processes causing magnetization transfer through the EPR spectrum, i.e. electron spin diffusion, nuclear relaxation, and slow rotational motions lead to different patterns and can be distinguished by recording 2D-ELDOR spectra as a function of temperature. In the specific system studied, the 2D-ELDOR spectra show the dominance of magnetization transfer between states with close molecular orientations but different nitrogen nuclear spin projections caused by flips of nuclear spins. The results are discussed in terms of dynamic processes in glasses.

  18. Synthesis and fluorescence properties of six fluorescein-nitroxide radical hybrid-compounds.

    PubMed

    Sato, Shingo; Endo, Susumu; Kurokawa, Yusuke; Yamaguchi, Masaki; Nagai, Akio; Ito, Tomohiro; Ogata, Tateaki

    2016-12-05

    Six fluorescein-nitroxide radical hybrid-compounds (2ab, 3ab, 4, and 5) were synthesized by the condensation of 5- or 6-carboxy-fluorescein and 4-amino-TEMPO (2ab), 5- or 6-aminofluorescein and 4-carboxy-TEMPO (3ab), and fluorescein and 4-carboxy-TEMPO (4), or by reaction of the 3-hydroxyl group of fluorescein with DPROXYL-3-ylmethyl methanesulfonate (5). Fluorescence intensities (around 520nm) after reduction of the radical increased to 1.43-, 1.38-, and 1.61-folds for 2a, 2b and 3b respectively; 3a alone exhibited a decrease in intensity on reduction. Since 4 was readily solvolyzed in PBS or even methanol to afford fluorescein and 4-carboxy-TEMPO, its fluorescence change could not be measured. Hybrid compound 5 containing an ether-linkage between the fluorescein phenol and 3-hydroxymethyl-DPROXYL hydroxyl centers, was stable and on reduction, showed a maximum increase (3.21-fold) in relative fluorescence intensity in PBS (pH5.0), despite its remarkably low absolute fluorescence intensity.

  19. Controlled Redox Chemistry at Cerium within a Tripodal Nitroxide Ligand Framework.

    PubMed

    Bogart, Justin A; Lippincott, Connor A; Carroll, Patrick J; Booth, Corwin H; Schelter, Eric J

    2015-12-01

    Ligand reorganization has been shown to have a profound effect on the outcome of cerium redox chemistry. Through the use of a tethered, tripodal, trianionic nitroxide ligand, [((2-tBuNOH)C6 H4 CH2 )3 N](3-) (TriNOx (3-) ), controlled redox chemistry at cerium was accomplished, and typically reactive complexes of tetravalent cerium were isolated. These included rare cationic complexes [Ce(TriNOx )thf][BAr(F) 4 ], in which Ar(F) =3,5-(CF3 )2 -C6 H3 , and [Ce(TriNOx )py][OTf]. A rare complete Ce-halide series, Ce(TriNOx )X, in which X=F(-) , Cl(-) , Br(-) , I(-) , was also synthesized. The solution chemistry of these complexes was explored through detailed solution-phase electrochemistry and (1) H NMR experiments and showed a unique shift in the ratio of species with inner- and outer-sphere anions with size of the anionic X(-) group. DFT calculations on the series of calculations corroborated the experimental findings.

  20. Effects of tempol and redox-cycling nitroxides in models of oxidative stress.

    PubMed

    Wilcox, Christopher S

    2010-05-01

    Tempol is a redox-cycling nitroxide that promotes the metabolism of many reactive oxygen species (ROS) and improves nitric oxide bioavailability. It has been studied extensively in animal models of oxidative stress. Tempol has been shown to preserve mitochondria against oxidative damage and improve tissue oxygenation. Tempol improved insulin responsiveness in models of diabetes mellitus and improved the dyslipidemia, reduced the weight gain and prevented diastolic dysfunction and heart failure in fat-fed models of the metabolic syndrome. Tempol protected many organs, including the heart and brain, from ischemia/reperfusion damage. Tempol prevented podocyte damage, glomerulosclerosis, proteinuria and progressive loss of renal function in models of salt and mineralocorticosteroid excess. It reduced brain or spinal cord damage after ischemia or trauma and exerted a spinal analgesic action. Tempol improved survival in several models of shock. It protected normal cells from radiation while maintaining radiation sensitivity of tumor cells. Its paradoxical pro-oxidant action in tumor cells accounted for a reduction in spontaneous tumor formation. Tempol was effective in some models of neurodegeneration. Thus, tempol has been effective in preventing several of the adverse consequences of oxidative stress and inflammation that underlie radiation damage and many of the diseases associated with aging. Indeed, tempol given from birth prolonged the life span of normal mice. However, presently tempol has been used only in human subjects as a topical agent to prevent radiation-induced alopecia.

  1. EELS Analysis of Nylon 6 Nanofibers Reinforced with Nitroxide-Functionalized Graphene Oxide.

    PubMed

    Leyva-Porras, César; Ornelas-Gutiérrez, C; Miki-Yoshida, M; Avila-Vega, Yazmín I; Macossay, Javier; Bonilla-Cruz, José

    2014-01-01

    A detailed analysis by transmission electron microscopy (TEM) and electron energy loss spectroscopy (EELS) of nitroxide-functionalized graphene oxide layers (GOFT) dispersed in Nylon 6 nanofibers is reported herein. The functionalization and exfoliation process of graphite oxide to GOFT was confirmed by TEM using electron diffraction patterns (EDP), wherein 1 to 4 graphene layers of GOFT were observed. The distribution and alignment of GOFT layers within a sample of Nylon 6 nanofiber reveals that GOFT platelets are mainly within the fiber, but some were partially protruding from it. Furthermore, Nylon 6 nanofibers exhibit an average diameter of 225 nm with several microns in length. GOFT platelets embedded into the fiber, the pristine fiber, and amorphous carbon were analyzed by EELS where each spectra [corresponding to the carbon edge (C-K)] exhibited changes in the fine structure, allowing a clear distinction between: i) GOFT single-layers, ii) Nylon-6 nanofibers, and iii) the carbon substrate. EELS analysis is presented here for the first time as a powerful tool to identify functionalized graphene single-layers (< 4 layers of GOFT) into a Nylon 6 nanofiber composite.

  2. EELS Analysis of Nylon 6 Nanofibers Reinforced with Nitroxide-Functionalized Graphene Oxide

    PubMed Central

    Leyva-Porras, César; Ornelas-Gutiérrez, C.; Miki-Yoshida, M.; Avila-Vega, Yazmín I.; Macossay, Javier; Bonilla-Cruz, José

    2014-01-01

    A detailed analysis by transmission electron microscopy (TEM) and electron energy loss spectroscopy (EELS) of nitroxide-functionalized graphene oxide layers (GOFT) dispersed in Nylon 6 nanofibers is reported herein. The functionalization and exfoliation process of graphite oxide to GOFT was confirmed by TEM using electron diffraction patterns (EDP), wherein 1 to 4 graphene layers of GOFT were observed. The distribution and alignment of GOFT layers within a sample of Nylon 6 nanofiber reveals that GOFT platelets are mainly within the fiber, but some were partially protruding from it. Furthermore, Nylon 6 nanofibers exhibit an average diameter of 225 nm with several microns in length. GOFT platelets embedded into the fiber, the pristine fiber, and amorphous carbon were analyzed by EELS where each spectra [corresponding to the carbon edge (C-K)] exhibited changes in the fine structure, allowing a clear distinction between: i) GOFT single-layers, ii) Nylon-6 nanofibers, and iii) the carbon substrate. EELS analysis is presented here for the first time as a powerful tool to identify functionalized graphene single-layers (< 4 layers of GOFT) into a Nylon 6 nanofiber composite. PMID:24634536

  3. Effects of tempol and redox-cycling nitroxides in models of oxidative stress

    PubMed Central

    Wilcox, Christopher S.

    2010-01-01

    Tempol is a redox cycling nitroxide that promotes the metabolism of many reactive oxygen species (ROS) and improves nitric oxide bioavailability. It has been studied extensively in animal models of oxidative stress. Tempol has been shown to preserve mitochondria against oxidative damage and improve tissue oxygenation. Tempol improved insulin responsiveness in models of diabetes mellitus and improved the dyslipidemia, reduced the weight gain and prevented diastolic dysfunction and heart failure in fat-fed models of the metabolic syndrome. Tempol protected many organs, including the heart and brain, from ischemia/reperfusion damage. Tempol prevented podocyte damage, glomerulosclerosis, proteinuria and progressive loss of renal function in models of salt and mineralocorticosteroid excess. It reduced brain or spinal cord damage after ischemia or trauma and exerted a spinal analgesic action. Tempol improved survival in several models of shock. It protected normal cells from radiation while maintaining radiation sensitivity of tumor cells. Its paradoxical pro-oxidant action in tumor cells accounted for a reduction in spontaneous tumor formation. Tempol was effective in some models of neurodegeneration. Thus, tempol has been effective in preventing several of the adverse consequences of oxidative stress and inflammation that underlie radiation damage and many of the diseases associated with aging. Indeed, tempol given from birth prolonged the life span of normal mice. However, presently tempol has been used only in human subjects as a topical agent to prevent radiation-induced alopecia. PMID:20153367

  4. Prediction of nitroxide spin label EPR spectra from MD trajectories: application to myoglobin.

    PubMed

    Kuprusevicius, Egidijus; White, Gaye; Oganesyan, Vasily S

    2011-01-01

    We report the prediction of motional EPR spectra of the metalloprotein sperm whale myoglobin spin labelled with nitroxide directly from Molecular Dynamics (MD) simulations at the atomistic scale. We show that an accurate simulation of EPR spectra can be achieved from a single MD trajectory which is in excellent agreement with experiment. Simulations have been carried out using a general method reported previously by us for the simulation of EPR spectra form single dynamical trajectories. Our calculations demonstrate the complex nature of the dynamics of a spin label which is a superposition of the fast librational motions around dihedral states, of slow conformational flips among different rotameric states and of the slow rotational diffusion of the protein itself. The MD-EPR methodology reported does not require any additional stochastic modelling using adjustable parameters and opens, for the first time, the prospect of the simulation of EPR spectra entirely from single MD trajectories. Such a technique not only simplifies the interpretation and analysis of EPR spectra but also opens the possibility, for example, of "computer engineering" of spin-labelled proteins with the desired properties prior to actual EPR experiment.

  5. Magnetic and structural properties of monoradicals and diradicals based on thienyl-substituted nitronyl nitroxide

    NASA Astrophysics Data System (ADS)

    Sugano, Tadashi; Blundell, Stephen J.; Hayes, William; Day, Peter; Mori, Hatsumi

    2010-06-01

    Magnetic properties of monoradicals and diradicals based on thienyl-substituted nitronyl nitroxide (NN) were investigated by measuring temperature and magnetic-field dependences of magnetization. The magnetic behavior of the monoradical 3-benzo[b]thienyl NN (3-BTHNN) is interpreted in terms of the quasi-2D ferromagnetic Heisenberg model with the coupling constant J/k=+0.16 K and J‧/k=+0.02 K, while the alternating antiferromagnetic (AFM) intermolecular interactions are observed in the monoradicals, 2- and 3-thienyl NN (2- and 3-THNN). In contrast, the magnetic behavior of the diradicals, 2,3- and 2,5-bis(NN)thiophene (2,3- and 2,5-THBNN), is explained by quite strong AFM intramolecular interactions between the two spin centers, each located on the two NN moieties. We discuss here the origin of the magnetic interactions observed in these radicals by considering their molecular and crystal structures determined by the X-ray diffraction techniques.

  6. Combined first-principles and thermodynamic approach to M -nitronyl nitroxide (M = Co, Mn) spin helices

    NASA Astrophysics Data System (ADS)

    Scarrozza, Marco; Vindigni, Alessandro; Barone, Paolo; Sessoli, Roberta; Picozzi, Silvia

    2015-04-01

    The properties of two molecular-based magnetic helices, composed of 3 d metal Co and Mn ions bridged by nitronyl nitroxide radicals, are investigated by density-functional calculations. Their peculiar and distinctive magnetic behavior is here elucidated by a thorough description of their magnetic, electronic, and anisotropy properties. Metal ions are antiferromagnetically coupled with the radicals, leading to a ferrimagnetically ordered ground state. A strong metal-radical exchange coupling is found, about 44 and 48 meV for Co and Mn helices, respectively. The latter have also relevant next-nearest-neighbor Mn-Mn antiferromagnetic interactions (of ˜6 meV). Co sites are characterized by noncollinear uniaxial anisotropies, whereas Mn sites are rather isotropic. A key result pertains to the Co helix: The microscopic picture resulting from density-functional calculations allows us to propose a spin Hamiltonian of increased complexity with respect to the commonly employed Ising Hamiltonian, suitable for the study of finite-temperature behavior, and that seems to clarify the puzzling scenario of multiple characteristic energy scales observed in experiments.

  7. Nuclear spin-lattice relaxation in nitroxide spin-label EPR.

    PubMed

    Marsh, Derek

    2016-11-01

    Nuclear relaxation is a sensitive monitor of rotational dynamics in spin-label EPR. It also contributes competing saturation transfer pathways in T1-exchange spectroscopy, and the determination of paramagnetic relaxation enhancement in site-directed spin labelling. A survey shows that the definition of nitrogen nuclear relaxation rate Wn commonly used in the CW-EPR literature for (14)N-nitroxyl spin labels is inconsistent with that currently adopted in time-resolved EPR measurements of saturation recovery. Redefinition of the normalised (14)N spin-lattice relaxation rate, b=Wn/(2We), preserves the expressions used for CW-EPR, whilst rendering them consistent with expressions for saturation recovery rates in pulsed EPR. Furthermore, values routinely quoted for nuclear relaxation times that are deduced from EPR spectral diffusion rates in (14)N-nitroxyl spin labels do not accord with conventional analysis of spin-lattice relaxation in this three-level system. Expressions for CW-saturation EPR with the revised definitions are summarised. Data on nitrogen nuclear spin-lattice relaxation times are compiled according to the three-level scheme for (14)N-relaxation: T1n=1/Wn. Results are compared and contrasted with those for the two-level (15)N-nitroxide system.

  8. Mitochondrial Dysfunction and Cell Death in Neurodegenerative Diseases through Nitroxidative Stress

    PubMed Central

    Akbar, Mohammed; Essa, Musthafa Mohamed; Daradkeh, Ghazi; Abdelmegeed, Mohamed A.; Choi, Youngshim; Mahmood, Lubna; Song, Byoung-Joon

    2016-01-01

    Mitochondria are important for providing cellular energy ATP through the oxidative phosphorylation pathway. They are also critical in regulating many cellular functions including the fatty acid oxidation, the metabolism of glutamate and urea, the anti-oxidant defense, and the apoptosis pathway. Mitochondria are an important source of reactive oxygen species leaked from the electron transport chain while they are susceptible to oxidative damage, leading to mitochondrial dysfunction and tissue injury. In fact, impaired mitochondrial function is commonly observed in many types of neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, alcoholic dementia, brain ischemia-reperfusion related injury, and others, although many of these neurological disorders have unique etiological factors. Mitochondrial dysfunction under many pathological conditions is likely to be promoted by increased nitroxidative stress, which can stimulate post-translational modifications (PTMs) of mitochondrial proteins and/or oxidative damage to mitochondrial DNA and lipids. Furthermore, recent studies have demonstrated that various antioxidants, including naturally occurring flavonoids and polyphenols as well as synthetic compounds, can block the formation of reactive oxygen and/or nitrogen species, and thus ultimately prevent the PTMs of many proteins with improved disease conditions. Therefore, the present review is aimed to describe the recent research developments in the molecular mechanisms for mitochondrial dysfunction and tissue injury in neurodegenerative diseases and discuss translational research opportunities. PMID:26883165

  9. Synthesis and fluorescence properties of six fluorescein-nitroxide radical hybrid-compounds

    NASA Astrophysics Data System (ADS)

    Sato, Shingo; Endo, Susumu; Kurokawa, Yusuke; Yamaguchi, Masaki; Nagai, Akio; Ito, Tomohiro; Ogata, Tateaki

    2016-12-01

    Six fluorescein-nitroxide radical hybrid-compounds (2ab, 3ab, 4, and 5) were synthesized by the condensation of 5- or 6-carboxy-fluorescein and 4-amino-TEMPO (2ab), 5- or 6-aminofluorescein and 4-carboxy-TEMPO (3ab), and fluorescein and 4-carboxy-TEMPO (4), or by reaction of the 3-hydroxyl group of fluorescein with DPROXYL-3-ylmethyl methanesulfonate (5). Fluorescence intensities (around 520 nm) after reduction of the radical increased to 1.43-, 1.38-, and 1.61-folds for 2a, 2b and 3b respectively; 3a alone exhibited a decrease in intensity on reduction. Since 4 was readily solvolyzed in PBS or even methanol to afford fluorescein and 4-carboxy-TEMPO, its fluorescence change could not be measured. Hybrid compound 5 containing an ether-linkage between the fluorescein phenol and 3-hydroxymethyl-DPROXYL hydroxyl centers, was stable and on reduction, showed a maximum increase (3.21-fold) in relative fluorescence intensity in PBS (pH 5.0), despite its remarkably low absolute fluorescence intensity.

  10. Nuclear spin-lattice relaxation in nitroxide spin-label EPR

    NASA Astrophysics Data System (ADS)

    Marsh, Derek

    2016-11-01

    Nuclear relaxation is a sensitive monitor of rotational dynamics in spin-label EPR. It also contributes competing saturation transfer pathways in T1-exchange spectroscopy, and the determination of paramagnetic relaxation enhancement in site-directed spin labelling. A survey shows that the definition of nitrogen nuclear relaxation rate Wn commonly used in the CW-EPR literature for 14N-nitroxyl spin labels is inconsistent with that currently adopted in time-resolved EPR measurements of saturation recovery. Redefinition of the normalised 14N spin-lattice relaxation rate, b = Wn/(2We), preserves the expressions used for CW-EPR, whilst rendering them consistent with expressions for saturation recovery rates in pulsed EPR. Furthermore, values routinely quoted for nuclear relaxation times that are deduced from EPR spectral diffusion rates in 14N-nitroxyl spin labels do not accord with conventional analysis of spin-lattice relaxation in this three-level system. Expressions for CW-saturation EPR with the revised definitions are summarised. Data on nitrogen nuclear spin-lattice relaxation times are compiled according to the three-level scheme for 14N-relaxation: T1n = 1/Wn. Results are compared and contrasted with those for the two-level 15N-nitroxide system.

  11. ESR study of molecular orientation and dynamics of nitronyl nitroxide radicals in CLPOT 1D nanochannels.

    PubMed

    Kobayashi, Hirokazu; Morinaga, Yuka; Fujimori, Etsuko; Asaji, Tetsuo

    2014-07-10

    New inclusion compounds (ICs) were prepared using the organic 1D nanochannels of 2,4,6-tris(4-chlorophenoxy)-1,3,5-triazine (CLPOT) as a nanosized template and nitronyl nitroxide (NN) radicals such as phenylnitronylnitroxide (PhNN) and p-nitrophenylnitronylnitroxide (p-NPNN). ESR measurements below 255 K for the CLPOT ICs diluted with spacer molecules gave rigid limit spectra similar to that for PhNN molecules in a glassy ethanol matrix at low temperature, which suggests isolation of the radical molecules. ESR measurements for them in the range of 290-400 K gave a modulated quintet ESR signal, which suggested uniaxial rotational diffusion of NN radicals in the nanochannels approximately around the principal y-axis of the g-tensors. In the ESR measurements to 430 K for the [(CLPOT)2-(p-NPNN)0.07] IC without spacers, the broader line width than the case in dilution was observed by inter-radical dipolar interaction. In every case, the rotational diffusion activation energies of NN radicals in the CLPOT nanochannels were several times larger than those of 2,2,6,6-tetramethyl-1-piperidinyloxyl (TEMPO) radical derivatives (4-X-TEMPO) in CLPOT nanochannels. This is expected due to the larger molecular size of NN radicals than 4-X-TEMPO or stronger interaction between NN radicals and the surrounding host or guest molecules.

  12. Synthesis of 14N and 15N-labeled trityl-nitroxide biradicals with strong spin-spin interaction and improved sensitivity to redox status and oxygen

    PubMed Central

    Liu, Yangping; Villamena, Frederick A.; Song, Yuguang; Sun, Jian; Rockenbauer, Antal

    2014-01-01

    Simultaneous evaluation redox status and oxygenation in biological systems is of great importance for the understanding of biological functions. Electron paramagnetic resonance spectroscopy coupled with the use of the nitroxide radicals have been an indispensable technique for this application but are still limited by its low oxygen sensitivity, and low EPR resolution in part due to the moderately broad EPR triplet and spin quenching through bioreduction. In this study, we showed that these drawbacks can be overcome through the use of trityl-nitroxide biradicals allowing for the simultaneous measurement of redox status and oxygenation. A new trityl-nitroxide biradical TNN14 composed of a pyrrolidinyl-nitroxide and a trityl, and its isotopically labeled 15N analogue TNN15 were synthesized and characterized. Both biradicals exhibited much stronger spin-spin interaction with J > 400 G than the previous synthesized trityl-nitroxide biradicals TN1 (~160 G) and TN2 (~52 G) with longer linker chain length. The enhanced stability of TNN14 was evaluated using ascorbate as reductant and the effect of different types of cyclodextrins on its stability in the presence of ascorbate was also investigated. Both biradicals are sensitive to redox status, and their corresponding trityl-hydroxylamines resulting from the reduction of the biradicals by ascorbate share the same oxygen sensitivity. Of note is that the 15N-labeled TNN15-H with an EPR doublet exhibits improved EPR signal amplitude as compared to TNN14-H with an EPR triplet. In addition, cyclic voltammetric studies verify the characteristic electrochemical behaviors of the trityl-nitroxide biradicals. PMID:21028905

  13. Site-specific insertion of nitroxide-spin labels into DNA probes by click chemistry for structural analyses by ELDOR spectroscopy.

    PubMed

    Flaender, M; Sicoli, G; Fontecave, Th; Mathis, G; Saint-Pierre, C; Boulard, Y; Gambarelli, S; Gasparutto, D

    2008-01-01

    A new approach is described for the insertion of nitroxide spin-labels at specific positions within DNA oligomers. The latter bioconjugaison strategy is based on a click chemistry 1,3-dipolar cycloaddition between a spin-labeling reagent, namely the 4-azido-TEMPO, and alkyne modified uridine-containing oligonucleotides. This highly efficient labeling method was applied for site-specific incorporation of two TEMPO units within a set of double-stranded DNA constructs. Then the determination of the inter-nitroxide distances was achieved by using a four-pulses DEER technique that successfully validates the new site-directed spin labeling strategy.

  14. ESR and HPLC-EC analysis of ethanol oxidation to 1-hydroxyethyl radical: rapid reduction and quantification of POBN and PBN nitroxides.

    PubMed

    Stoyanovsky, D A; Cederbaum, A I

    1998-09-01

    Extensive ESR spin-trapping studies have shown that ethanol is oxidized to 1-hydroxyethyl radical (HER) by rat and deer mice liver microsomal systems. The ESR detection of POBN/HER nitroxide in bile, and formation of antibodies, which recognize HER adducts in alcoholics, suggest that HER is produced in vivo. In liver, where ethanol is primarily metabolized, only traces of PBN/HER nitroxide are documented. One limitation of the ESR spin-trapping technique, however, is that the nitroxides formed in the presence of cellular reductants can be metabolized to the corresponding ESR "silent" hydroxylamines. Ascorbate and NADPH plus liver microsomes were found to reduce rapidly both POBN/HER and PBN/HER nitroxides to their ESR "silent" hydroxylamine derivatives. An HPLC method with electrochemical detection was developed for the detection and quantification of both POBN/HER and PBN/HER nitroxides, as well as their hydroxylamines. Both the diastereomers of the POBN/HER nitroxide and hydroxylamine can be detected, as can both isomers of the PBN/HER nitroxide, and it is estimated that the sensitivity of the HPLC procedure is in the nM range when using EC detection. The hydroxylamines are stable in ethanol, while pH-dependent auto-oxidation occurs in aqueous buffers. Some of the characteristics associated with HER formation by microsomes as detected with ESR (e.g., sensitivity to SOD and catalase, increase after induction of CYP2E1) are reproduced with the HPLC method. By quantification of the POBN/HER hydroxylamine, the NADPH-dependent rates of HER formation by microsomes from pyrazole-treated rats are estimated to be about 1-1.5 nmol HER per min per mg microsomal protein. This rate is less, as compared to the two electron-dependent rate of acetaldehyde formation by these microsomes, about 10-15 nmol per min per mg protein. Thus, at first approximation, the one electron-dependent rate of ethanol oxidation is about 10% the two electron-dependent rate by isolated microsomes. The

  15. Astrocytes expressing mutant SOD1 and TDP43 trigger motoneuron death that is mediated via sodium channels and nitroxidative stress

    PubMed Central

    Rojas, Fabiola; Cortes, Nicole; Abarzua, Sebastian; Dyrda, Agnieszka; van Zundert, Brigitte

    2013-01-01

    Amyotrophic lateral sclerosis (ALS) is a fatal paralytic disorder caused by dysfunction and degeneration of motor neurons. Multiple disease-causing mutations, including in the genes for SOD1 and TDP-43, have been identified in ALS. Astrocytes expressing mutant SOD1 are strongly implicated in the pathogenesis of ALS: we have shown that media conditioned by astrocytes carrying mutant SOD1G93A contains toxic factor(s) that kill motoneurons by activating voltage-sensitive sodium (Nav) channels. In contrast, a recent study suggests that astrocytes expressing mutated TDP43 contribute to ALS pathology, but do so via cell-autonomous processes and lack non-cell-autonomous toxicity. Here we investigate whether astrocytes that express diverse ALS-causing mutations release toxic factor(s) that induce motoneuron death, and if so, whether they do so via a common pathogenic pathway. We exposed primary cultures of wild-type spinal cord cells to conditioned medium derived from astrocytes (ACM) that express SOD1 (ACM-SOD1G93A and ACM-SOD1G86R) or TDP43 (ACM-TDP43A315T) mutants; we show that such exposure rapidly (within 30–60 min) increases dichlorofluorescein (DCF) fluorescence (indicative of nitroxidative stress) and leads to extensive motoneuron-specific death within a few days. Co-application of the diverse ACMs with anti-oxidants Trolox or esculetin (but not with resveratrol) strongly improves motoneuron survival. We also find that co-incubation of the cultures in the ACMs with Nav channel blockers (including mexiletine, spermidine, or riluzole) prevents both intracellular nitroxidative stress and motoneuron death. Together, our data document that two completely unrelated ALS models lead to the death of motoneuron via non-cell-autonomous processes, and show that astrocytes expressing mutations in SOD1 and TDP43 trigger such cell death through a common pathogenic pathway that involves nitroxidative stress, induced at least in part by Nav channel activity. PMID:24570655

  16. Novel multi-responsive P2VP-block-PNIPAAm block copolymers via nitroxide-mediated radical polymerization

    PubMed Central

    Corten, Cathrin; Kretschmer, Katja

    2010-01-01

    Summary Linear soluble multi-responsive block copolymers are able to form so called schizophrenic micelles in aqueous solution. Here, such polymers are prepared via nitroxide-mediated radical polymerization (NMRP). In a first step nitroxide-terminated poly(2-vinylpyridine) (P2VP) was prepared with different molecular weights and narrow molecular weight distributions. The best reaction conditions, optimized by kinetic studies, were bulk polymerization at 110 °C. Using P2VP as a macroinitiator, the synthesis of new soluble linear block copolymers of P2VP and poly(N-isopropylacrylamide) (PNIPAAm) (P2VP-block-PNIPAAm) was possible. The nitroxide terminated polymers were characterized by nuclear magnetic resonance (NMR) spectroscopy, size exclusion chromatography (SEC) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Thermal properties were investigated by the differential scanning calorimetry (DSC). Block copolymers showed pH- and temperature-responsive solubility in aqueous media. By increasing the P2VP content, the phase transition temperature shifted to lower temperatures (e.g. 26 °C for P2VP114-block-PNIPAAm180). Depending on the resulting block length, temperature and pH value of aqueous solution, the block copolymers form so called schizophrenic micelles. The hydrodynamic radius R h of these micelles associated with pH values and temperature was analyzed by dynamic light scattering (DLS). Such kind of block copolymers has potential for many applications, such as controlled drug delivery systems. PMID:20978627

  17. [Application of ESR imaging technique in studying of skin-penetration properties of nitroxide free radical].

    PubMed

    Wu, Ke; Zheng, Yingguang; Cong, Jianbo; Zhang, Qingjun; Wang, Changzhen; Xian, Hong; Sun, Cunpu

    2008-08-01

    A set of L-band electron spin resonance imaging (ESRI) equipment suitable for biological species was developed and an ESRI experiment model for viable skin samples was established. The mechanic process of nitroxide free radical TEMPO (2,2, 6, 6-tetramethyl-1-piperidinyloxy) penetrating through skin sample and the spin density distribution of TEMPO after it interacted with skin sample were detected by the developed ESRI method. Skin samples were extracted from mice back. The experimental samples were prepared by cutting the skin pieces into square shape of 2 x 2 cm2 and then the samples were divided into three groups by treating them with three different methods: Method A, simple treatment by simply cutting the hair; method B, 8% Na2S depilation treatment for 10 min; method C, 8% Na2S depilation and then 5% pancreatic digestion treatment for 2 hours. The liposoluble solvent DMSO (dimethyl sulfoxide) and distilled water were used as two kinds of solvent for the TEMPO liquor. The results indicated that the skin-penetration properties of TEMPO were significantly different among samples treated with different methods and the surface cornifin of skin offered remarkable resistance to TEMPO. The TEMPO liquor of water could hardly penetrate through skins, whereas about 20%-30% of the original TEMPO compounds that solved in liposoluble solvent DMSO could penetrate through the skin sample treated with method C after 16 hours of interaction. Furthermore, the penetration rate of TEMPO through the skin tissue was a strong time dependent process. The preliminary application results suggested that ESRI technique could provide an effective and applicable method for dynamically researching skin-penetration properties of some special kinds of materials such as paramagnetic compounds.

  18. Quenching of the perylene fluorophore by stable nitroxide radical-containing macromolecules.

    PubMed

    Hughes, Barbara K; Braunecker, Wade A; Ferguson, Andrew J; Kemper, Travis W; Larsen, Ross E; Gennett, Thomas

    2014-10-30

    Stable nitroxide radical bearing organic polymer materials are attracting much attention for their application as next generation energy storage materials. A greater understanding of the inherent charge transfer mechanisms in such systems will ultimately be paramount to further advancements in the understanding of both intrafilm and interfacial ion- and electron-transfer reactions. This work is focused on advancing the fundamental understanding of these dynamic charge transfer properties by exploiting the fact that these species are efficient fluorescence quenchers. We systematically incorporated fluorescent perylene dyes into solutions containing the 2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPO) radical and controlled their interaction by binding the TEMPO moiety into macromolecules with varying morphologies (e.g., chain length, density of radical pendant groups). In the case of the model compound, 4-oxo-TEMPO, quenching of the perylene excited state was found to be dominated by a dynamic (collisional) process, with a contribution from an apparent static process that is described by an ∼2 nm quenching sphere of action. When we incorporated the TEMPO unit into a macromolecule, the quenching behavior was altered significantly. The results can be described by using two models: (A) a collisional quenching process that becomes less efficient, presumably due to a reduction in the diffusion constant of the quenching entity, with a quenching sphere of action similar to 4-oxo-TEMPO or (B) a collisional quenching process that becomes more efficient as the radius of interaction grows larger with increasing oligomer length. This is the first study that definitively illustrates that fluorophore quenching by a polymer system cannot be explained using merely a classical Stern-Volmer approach but rather necessitates a more complex model.

  19. Analysis of the Magnetic Exchange Interactions in Yttrium(III) Complexes Containing Nitronyl Nitroxide Radicals.

    PubMed

    Jung, Julie; Puget, Marin; Cador, Olivier; Bernot, Kevin; Calzado, Carmen J; Le Guennic, Boris

    2017-06-19

    We report a combined theoretical and experimental investigation of the exchange interactions governing the magnetic behavior of a series of nitronyl nitroxide (NIT)-based Y(III) complexes, i.e., Y(hfac)3(NIT-R)2 with R = PhOPh (1), PhOEt (2), and PhOMe (3a, 3b). Even though some of these complexes or their Dy(III) parents were previously described in the literature [ Zhao et al. Transition Met. Chem. 2006 , 31 , 593 ; Bernot et al. J. Am. Chem. Soc. 2009 , 131 , 5573 ], their synthesis procedure as well as their structural and magnetic properties were completely reconsidered. Depending on the nature of R and the crystallization conditions, Y(hfac)3(NIT-R)2 units can be organized as supramolecular dimers or linear or orthogonal chains. Such structural diversity within the series induces extremely different magnetic behaviors. The observed behaviors are rationalized by state-of-the-art wave function-based quantum-chemical approaches (CASSCF/DDCI) that demonstrate the existence of not only effective intramolecular interactions between the NIT-R radical ligands of an isolated Y(hfac)3(NIT-R)2 molecule but also intermolecular interactions between NIT-R moieties belonging to different Y(hfac)3(NIT-R)2 units. These results are supported by the use of spin Hamiltonian models going beyond the basic Bleaney-Bowers formalism to properly fit the experimental magnetic data. Finally, the microscopic mechanisms behind the evidenced intramolecular exchange interactions are elucidated through the inspection of the calculated wave functions. In particular, whereas the role of Y orbitals was already proposed, we herein demonstrate the contribution of the hfac(-) ancillary ligands in mediating the magnetic interactions between the NIT radicals.

  20. Anion dependent redox changes in iron bis-terdentate nitroxide {NNO} chelates.

    PubMed

    Gass, Ian A; Gartshore, Christopher J; Lupton, David W; Moubaraki, Boujemaa; Nafady, Ayman; Bond, Alan M; Boas, John F; Cashion, John D; Milsmann, Carsten; Wieghardt, Karl; Murray, Keith S

    2011-04-04

    The reaction of [Fe(II)(BF(4))(2)]·6H(2)O with the nitroxide radical, 4,4-dimethyl-2,2-di(2-pyridyl) oxazolidine-N-oxide (L(•)), produces the mononuclear transition metal complex [Fe(II)(L(•))(2)](BF(4))(2) (1) which has been investigated using temperature dependent susceptibility, Mössbauer spectroscopy, electrochemistry, density functional theory (DFT) calculations, and X-ray structure analysis. Single crystal X-ray diffraction analysis and Mössbauer measurements reveal an octahedral low spin Fe(2+) environment where the pyridyl donors from L(•) coordinate equatorially while the oxygen containing the radical from L(•) coordinates axially forming a linear O(•)··Fe(II)··O(•) arrangement. Magnetic susceptibility measurements show a strong radical-radical intramolecular antiferromagnetic interaction mediated by the diamagnetic Fe(2+) center. This is supported by DFT calculations which show a mutual spatial overlap of 0.24 and a spin density population analysis which highlights the antiparallel spin alignment between the two ligands. Similarly the monocationic complex [Fe(III)(L(-))(2)](BPh(4))·0.5H(2)O (2) has been fully characterized with Fe-ligand and N-O bond length changes in the X-ray structure analysis, magnetic measurements revealing a Curie-like S = 1/2 ground state, electron paramagnetic resonance (EPR) spectra, DFT calculations, and electrochemistry measurements all consistent with assignment of Fe in the (III) state and both ligands in the L(-) form. 2 is formed by a rare, reductively induced oxidation of the Fe center, and all physical data are self-consistent. The electrochemical studies were undertaken for both 1 and 2, thus allowing common Fe-ligand redox intermediates to be identified and the results interpreted in terms of square reaction schemes.

  1. Spin-Parity Behavior in the Exchange-Coupled Lanthanoid-Nitroxide Molecular Magnets

    NASA Astrophysics Data System (ADS)

    Ishida, T.

    2017-05-01

    To develop lanthanoid-based magnetic materials and relevant devices, reliable prescriptions for molecular/crystal design have long been desired. Ln3+-ion dependence on the molecular magnetism was investigated in the isomorphous series [Ln(hfac)3(2pyNO)] (Ln = Tb, Dy, Ho, Er), where 2pyNO stands for tert-butyl 2-pyridyl nitroxide as a paramagnetic ligand, and hfac for 1,1,1,5,5,5-hexafluoropentane-2,4-dionate. The slow magnetization reversal was evaluated as an indication of single-molecule magnets (SMMs) by out-of-phase ac magnetic susceptibility χ”. Whereas the Tb3+ (4f8) and Ho3+ (4f10) derivatives exhibited frequency-dependent χ”, practically null χ” was recorded for the Dy3+(4f9) and Er3+ (4f11) derivatives. As for another series with Ln/radical = 1/2, [Ln(hfac)3(TEMPO)2] complexes were prepared (Ln = Tb, Dy, Ho, Er, Tm; TEMPO = 2,2,6,6-tetramethylpiperidin-1-oxyl). The Dy3+ and Er3+ derivatives showed appreciable χ”, but the Tb3+, Ho3+, and Tm3+ derivatives did not. Thus, the S = 1/2 paramagnetic ligands play a role of a spin-parity switch to regulate whether the compound behaves as an SMM. In the strongly exchange-coupled regime owing to the direct radical coordination bond, the whole molecular electron counting may provide a useful criterion to predict Kramers molecules and accordingly to explore potential SMM candidates.

  2. Suppression of B-Raf(V600E) melanoma cell survival by targeting mitochondria using triphenyl-phosphonium-conjugated nitroxide or ubiquinone.

    PubMed

    Hong, Seung-Keun; Starenki, Dmytro; Wu, Pui-Kei; Park, Jong-In

    2017-02-01

    Most BRAF-mutated melanomas initially responsive to the FDA-approved inhibitors preferentially targeting B-Raf mutated in Val600 residue eventually relapse, requiring additional therapeutic modalities. Recent studies report the significance of metabolic reprograming in mitochondria for maintenance of BRAF-mutated melanomas and for development of their drug resistance to B-Raf inhibitors, providing a rationale for targeting mitochondria as a potential therapeutic strategy for melanoma. We therefore determined whether mitochondria-targeted metabolism-interfering agents can effectively suppress human B-Raf(V600E) melanoma cell lines and their dabrafenib/PLX4032-resistant progenies using mitochondria-targeted carboxy-proxyl (Mito-CP) and ubiquinone (Mito-Q). These agents exhibited comparable efficacy to PLX4032 in suppressing SK-MEL28, A375, and RPMI-7951 cells in vitro. As determined in SK-MEL28 and A375 cells, Mito-CP induced apoptotic cell death mediated by mitochondrial membrane depolarization and subsequent oxidative stress, which PLX4032 could not induce. Of note, Mito-CP also effectively suppressed PLX4032-resistant progenies of SK-MEL28 and A375. Moreover, when orally administered, Mito-CP suppressed SK-MEL28 xenografts in mice as effectively as PLX4032 without serious adverse effects. These data demonstrate that mitochondria-targeted agents have therapeutic potential to effectively suppress BRAF-mutated melanomas via an effect(s) distinct from those of B-Raf inhibitors.

  3. Synthesis and evaluation of nitroxide-based oligoradicals for low-temperature dynamic nuclear polarization in solid state NMR.

    PubMed

    Yau, Wai-Ming; Thurber, Kent R; Tycko, Robert

    2014-07-01

    We describe the synthesis of new nitroxide-based biradical, triradical, and tetraradical compounds and the evaluation of their performance as paramagnetic dopants in dynamic nuclear polarization (DNP) experiments in solid state nuclear magnetic resonance (NMR) spectroscopy with magic-angle spinning (MAS). Under our experimental conditions, which include temperatures in the 25-30 K range, a 9.4 T magnetic field, MAS frequencies of 6.2-6.8 kHz, and microwave irradiation at 264.0 GHz from a 800 mW extended interaction oscillator source, the most effective compounds are triradicals that are related to the previously-described compound DOTOPA-TEMPO (see Thurber et al., 2010), but have improved solubility in glycerol/water solvent near neutral pH. Using these compounds at 30 mM total nitroxide concentration, we observe DNP enhancement factors of 92-128 for cross-polarized (13)C NMR signals from (15)N,(13)C-labeled melittin in partially protonated glycerol/water, and build-up times of 2.6-3.8s for (1)H spin polarizations. Net sensitivity enhancements with biradical and tetraradical dopants, taking into account absolute (13)C NMR signal amplitudes and build-up times, are approximately 2-4 times lower than with the best triradicals. Published by Elsevier Inc.

  4. Dipolar Coupling between Nitroxide Spin Labels: The Development and Application of a Tether-in-a-Cone Model

    PubMed Central

    Hustedt, Eric J.; Stein, Richard A.; Sethaphong, Latsavongsakda; Brandon, Suzanne; Zhou, Zheng; DeSensi, Susan C.

    2006-01-01

    A tether-in-a-cone model is developed for the simulation of electron paramagnetic resonance spectra of dipolar coupled nitroxide spin labels attached to tethers statically disordered within cones of variable halfwidth. In this model, the nitroxides adopt a range of interprobe distances and orientations. The aim is to develop tools for determining both the distance distribution and the relative orientation of the labels from experimental spectra. Simulations demonstrate the sensitivity of electron paramagnetic resonance spectra to the orientation of the cones as a function of cone halfwidth and other parameters. For small cone halfwidths (<∼40°), simulated spectra are strongly dependent on the relative orientation of the cones. For larger cone halfwidths, spectra become independent of cone orientation. Tether-in-a-cone model simulations are analyzed using a convolution approach based on Fourier transforms. Spectra obtained by the Fourier convolution method more closely fit the tether-in-a-cone simulations as the halfwidth of the cone increases. The Fourier convolution method gives a reasonable estimate of the correct average distance, though the distance distribution obtained can be significantly distorted. Finally, the tether-in-a-cone model is successfully used to analyze experimental spectra from T4 lysozyme. These results demonstrate the utility of the model and highlight directions for further development. PMID:16214868

  5. Amelioration of Radiation Esophagitis by Orally Administered p53/Mdm2/Mdm4 Inhibitor (BEB55) or GS-Nitroxide

    PubMed Central

    KIM, HYUN; BERNARD, MARK E.; EPPERLY, MICHAEL W.; SHEN, HONGMEI; AMOSCATO, ANDREW; DIXON, TRACY M.; DOEMLING, ALEXANDER S.; LI, SONG; GAO, XIANG; WIPF, PETER; WANG, HONG; ZHANG, XICHEN; KAGAN, VALERIAN E.; GREENBERGER, JOEL S.

    2012-01-01

    Background/Aim Esophagitis is a significant toxicity of radiation therapy for lung cancer. In this study, reduction of irradiation esophagitis in mice, by orally administered p53/Mdm2/Mdm4 inhibitor, BEB55, or the GS-nitroxide, JP4-039, was evaluated. Materials and Methods BEB55 or JP4-039 in F15 (liposomal) formulation was administered intraesophageally to C57BL/6 mice prior to thoracic irradiation of 29 Gy × 1 or 11.5 Gy × 4 thoracic irradiation. Progenitor cells were sorted from excised esophagus, and nitroxide was quantified, by electron paramagnetic resonance (EPR). Mice with Lewis lung carcinoma (3LL) orthotopic lung tumors were treated with BEB55 or JP4-039 prior to 20 Gy to determine if the drugs would protect the tumor cells from radiation. Results Intraesophageal BEB55 and JP4-039 compared to formulation alone increased survival after single fraction (p=0.0209 and 0.0384, respectively) and four fraction thoracic irradiation (p=0.0241 and 0.0388, respectively). JP4-039 was detected in esophagus, liver, bone marrow, and orthotopic Lewis lung carcinoma (3LL) tumor. There was no significant radiation protection of lung tumors by BEB55 or JP4-039 compared to formulation only as assessed by survival (p=0.3021 and 0.3693, respectively). Thus, BEB55 and JP4-039 safely ameliorate radiation esophagitis in mice. PMID:22021675

  6. Intraesophageal Administration of GS-Nitroxide (JP4-039) Protects Against Ionizing Irradiation-induced Esophagitis

    PubMed Central

    EPPERLY, MICHAEL W.; GOFF, JULIE P.; LI, SONG; GAO, XIANG; WIPF, PETER; DIXON, TRACY; WANG, HONG; FRANICOLA, DARCY; SHEN, HONGMEI; RWIGEMA, JEAN-CLAUDE M.; KAGAN, VALERIAN; BERNARD, MARK; GREENBERGER, JOEL S.

    2012-01-01

    Background/Aim This study evaluated esophageal radioprotection by the Gramicidin S (GS) derived-nitroxide, JP4-039, a mitochondrial targeting peptide-isostere covalently-linked to 4-amino-Tempo, delivered in a novel swallowed oil-based (F15) formulation. Materials and Methods C57BL/6HNsd female mice received intraesophageal F15 formulation containing JP4-039 (4 mg/ml in 100 μl volumes) 10 minutes before 28 or 29 Gy upper body irradiation compared to MnSOD-PL (100 μl containing 100 μg plasmid) 24 hours prior to irradiation. Subgroups received 1×107 C57BL/6HNsd, GFP+ male bone marrow cells intravenously 5 days after irradiation. Results JP4-039/F15 or MnSOD-PL increased survival compared to irradiated controls (p<0.0001 for either). Marrow injection further increased survival (p=0.0462 and 0.0351, respectively). Esophagi removed at 1, 3, 7, 14, 24, or 60 days showed bone marrow-derived cells in the esophagi. Conclusion Intraesophageal GS-nitroxide radioprotection is mediated primarily through recovery of endogenous esophageal progenitor cells. PMID:21164038

  7. Two-dimensional electron paramagnetic resonance spectroscopy of nitroxides: Elucidation of restricted molecular motions in glassy solids

    NASA Astrophysics Data System (ADS)

    Dubinskii, Alexander A.; Maresch, Günter G.; Spiess, Hans-Wolfgang

    1994-02-01

    The combination of concepts of two-dimensional (2D) spectroscopy with the well-known field step electron-electron double resonance (ELDOR) method offers a practical route to recording 2D ELDOR spectra covering the full spectral range needed for electron paramagnetic resonance (EPR) of nitroxide spin labels in the solid state. The 2D ELDOR pattern provides information about molecular reorientation measured in real time, the anisotropies of electron phase, and electron spin-lattice relaxation as well as nuclear spin-lattice relaxation all of which are connected with the detailed geometry of the molecular reorientation. Thus, in 2D ELDOR the same electron spin probes the motional behavior over a wide range of correlation times from 10-4 to 10-12 s. An efficient algorithm for simulating 2D ELDOR spectra is derived, based on analytical solutions of the spin relaxation behavior for small-angle fluctuations and offers a means of quantitatively analyzing experimental data. As an example, the motion of nitroxide spin labels in a liquid-crystalline side-group polymer well below its glass transition is determined as a β-relaxation process with a mean angular amplitude of 5° and a distribution of correlation times with a mean correlation time of 0.9×10-10 s and a width of 2.5 decades.

  8. Synthesis and evaluation of nitroxide-based oligoradicals for low-temperature dynamic nuclear polarization in solid state NMR

    PubMed Central

    Yau, Wai-Ming; Thurber, Kent R.; Tycko, Robert

    2014-01-01

    We describe the synthesis of new nitroxide-based biradical, triradical, and tetraradical compounds and the evaluation of their performance as paramagnetic dopants in dynamic nuclear polarization (DNP) experiments in solid state nuclear magnetic resonance (NMR) spectroscopy with magic-angle spinning (MAS). Under our experimental conditions, which include temperatures in the 25–30 K range, a 9.4 T magnetic field, MAS frequencies of 6.2–6.8 kHz, and microwave irradiation at 264.0 GHz from a 800 mW extended interaction oscillator source, the most effective compounds are triradicals that are related to the previously-described compound DOTOPA-TEMPO (see Thurber et al., 2010), but have improved solubility in glycerol/water solvent near neutral pH. Using these compounds at 30 mM total nitroxide concentration, we observe DNP enhancement factors of 92–128 for cross-polarized 13C NMR signals from 15N,13C-labeled melittin in partially protonated glycerol/water, and build-up times of 2.6–3.8 s for 1H spin polarizations. Net sensitivity enhancements with biradical and tetraradical dopants, taking into account absolute 13C NMR signal amplitudes and build-up times, are approximately 2–4 times lower than with the best triradicals. PMID:24887201

  9. Nitroxidative chemistry interferes with fluorescent probe chemistry: implications for nitric oxide detection using 2,3-diaminonaphthalene.

    PubMed

    Hu, Teh-Min; Chiu, Shih-Jiuan; Hsu, Yu-Ming

    2014-08-22

    Simultaneous production of nitric oxide (NO) and superoxide generates peroxynitrite and causes nitroxidative stress. The fluorometric method for NO detection is based on the formation of a fluorescent product from the reaction of a nonfluorescent probe molecule with NO-derived nitrosating species. Here, we present an example of how nitroxidative chemistry could interact with fluorescent probe chemistry. 2,3-Naphthotriazole (NAT) is the NO-derived fluorescent product of 2,3-diaminonaphthalene (DAN), a commonly used NO-detecting molecule. We show that NO/superoxide cogeneration, and particularly peroxynitrite, mediates the chemical decomposition of NAT. Moreover, the extent of NAT decomposition depends on the relative fluxes of NO and superoxide; the maximum effect being reached at almost equivalent generation rates for both radicals. The rate constant for the reaction of NAT with peroxynitrite was determined to be 2.2×10(3)M(-1)s(-1). Further, various peroxynitrite scavengers were shown to effectively inhibit NO/superoxide- and peroxynitrite-mediated decomposition of NAT. Taken together, the present study suggests that the interference of a fluorometric NO assay can be originated from the interaction between the final fluorescent product and the formed reactive nitrogen and oxygen species.

  10. Perturbation of nuclear spin polarizations in solid state NMR of nitroxide-doped samples by magic-angle spinning without microwaves.

    PubMed

    Thurber, Kent R; Tycko, Robert

    2014-05-14

    We report solid state (13)C and (1)H nuclear magnetic resonance (NMR) experiments with magic-angle spinning (MAS) on frozen solutions containing nitroxide-based paramagnetic dopants that indicate significant perturbations of nuclear spin polarizations without microwave irradiation. At temperatures near 25 K, (1)H and cross-polarized (13)C NMR signals from (15)N,(13)C-labeled L-alanine in trinitroxide-doped glycerol/water are reduced by factors as large as six compared to signals from samples without nitroxide doping. Without MAS or at temperatures near 100 K, differences between signals with and without nitroxide doping are much smaller. We attribute most of the reduction of NMR signals under MAS near 25 K to nuclear spin depolarization through the cross-effect dynamic nuclear polarization mechanism, in which three-spin flips drive nuclear polarizations toward equilibrium with spin polarization differences between electron pairs. When T1e is sufficiently long relative to the MAS rotation period, the distribution of electron spin polarization across the nitroxide electron paramagnetic resonance lineshape can be very different from the corresponding distribution in a static sample at thermal equilibrium, leading to the observed effects. We describe three-spin and 3000-spin calculations that qualitatively reproduce the experimental observations.

  11. W-band orientation selective DEER measurements on a Gd3+/nitroxide mixed-labeled protein dimer with a dual mode cavity

    NASA Astrophysics Data System (ADS)

    Kaminker, Ilia; Tkach, Igor; Manukovsky, Nurit; Huber, Thomas; Yagi, Hiromasa; Otting, Gottfried; Bennati, Marina; Goldfarb, Daniella

    2013-02-01

    Double electron-electron resonance (DEER) at W-band (95 GHz) was applied to measure the distance between a pair of nitroxide and Gd3+ chelate spin labels, about 6 nm apart, in a homodimer of the protein ERp29. While high-field DEER measurements on systems with such mixed labels can be highly attractive in terms of sensitivity and the potential to access long distances, a major difficulty arises from the large frequency spacing (about 700 MHz) between the narrow, intense signal of the Gd3+ central transition and the nitroxide signal. This is particularly problematic when using standard single-mode cavities. Here we show that a novel dual-mode cavity that matches this large frequency separation dramatically increases the sensitivity of DEER measurements, allowing evolution times as long as 12 μs in a protein. This opens the possibility of accessing distances of 8 nm and longer. In addition, orientation selection can be resolved and analyzed, thus providing additional structural information. In the case of W-band DEER on a Gd3+-nitroxide pair, only two angles and their distributions have to be determined, which is a much simpler problem to solve than the five angles and their distributions associated with two nitroxide spin labels.

  12. W-band orientation selective DEER measurements on a Gd3+/nitroxide mixed-labeled protein dimer with a dual mode cavity.

    PubMed

    Kaminker, Ilia; Tkach, Igor; Manukovsky, Nurit; Huber, Thomas; Yagi, Hiromasa; Otting, Gottfried; Bennati, Marina; Goldfarb, Daniella

    2013-02-01

    Double electron-electron resonance (DEER) at W-band (95 GHz) was applied to measure the distance between a pair of nitroxide and Gd(3+) chelate spin labels, about 6 nm apart, in a homodimer of the protein ERp29. While high-field DEER measurements on systems with such mixed labels can be highly attractive in terms of sensitivity and the potential to access long distances, a major difficulty arises from the large frequency spacing (about 700 MHz) between the narrow, intense signal of the Gd(3+) central transition and the nitroxide signal. This is particularly problematic when using standard single-mode cavities. Here we show that a novel dual-mode cavity that matches this large frequency separation dramatically increases the sensitivity of DEER measurements, allowing evolution times as long as 12 μs in a protein. This opens the possibility of accessing distances of 8 nm and longer. In addition, orientation selection can be resolved and analyzed, thus providing additional structural information. In the case of W-band DEER on a Gd(3+)-nitroxide pair, only two angles and their distributions have to be determined, which is a much simpler problem to solve than the five angles and their distributions associated with two nitroxide spin labels. Copyright © 2012 Elsevier Inc. All rights reserved.

  13. Perturbation of nuclear spin polarizations in solid state NMR of nitroxide-doped samples by magic-angle spinning without microwaves

    SciTech Connect

    Thurber, Kent R. Tycko, Robert

    2014-05-14

    We report solid state {sup 13}C and {sup 1}H nuclear magnetic resonance (NMR) experiments with magic-angle spinning (MAS) on frozen solutions containing nitroxide-based paramagnetic dopants that indicate significant perturbations of nuclear spin polarizations without microwave irradiation. At temperatures near 25 K, {sup 1}H and cross-polarized {sup 13}C NMR signals from {sup 15}N,{sup 13}C-labeled L-alanine in trinitroxide-doped glycerol/water are reduced by factors as large as six compared to signals from samples without nitroxide doping. Without MAS or at temperatures near 100 K, differences between signals with and without nitroxide doping are much smaller. We attribute most of the reduction of NMR signals under MAS near 25 K to nuclear spin depolarization through the cross-effect dynamic nuclear polarization mechanism, in which three-spin flips drive nuclear polarizations toward equilibrium with spin polarization differences between electron pairs. When T{sub 1e} is sufficiently long relative to the MAS rotation period, the distribution of electron spin polarization across the nitroxide electron paramagnetic resonance lineshape can be very different from the corresponding distribution in a static sample at thermal equilibrium, leading to the observed effects. We describe three-spin and 3000-spin calculations that qualitatively reproduce the experimental observations.

  14. Synthesis and spectral properties of polymethine-cyanine dye-nitroxide radical hybrid compounds for use as fluorescence probes to monitor reducing species and radicals

    NASA Astrophysics Data System (ADS)

    Sato, Shingo; Tsunoda, Minoru; Suzuki, Minoru; Kutsuna, Masahiro; Takido-uchi, Kiyomi; Shindo, Mitsuru; Mizuguchi, Hitoshi; Obara, Heitaro; Ohya, Hiroaki

    2009-01-01

    Various hybrid compounds comprised of two types of nitroxide radicals and either a pentamethine (Cy5) or trimethine cyanine (Cy3) were synthesized. The nitroxide radicals were linked either via an ester-bond to one or two N-alkyl carboxyl-terminated groups of Cy5, or via two amido-bonds (aminocarbonyl or carbonylamino group) to the 5-position of the indolenine moieties of Cy5 and Cy3. Changes in fluorescence and ESR intensities of the hybrid compounds were measured before and after addition of Na ascorbate in PBS (pH 7.0) to reduce the radicals. Among the hybrid compounds synthesized, those that linked the nitroxide radicals via an aminocarbonyl residue at the 5-position of the indolenine moieties on Cy5 and Cy3 exhibited a 1.8- and 5.1-fold increase in fluorescence intensity with the reduction of the nitroxide segment by the addition of Na ascorbate, respectively. In contrast, fluorescence intensity was not enhanced in the other hybrid compounds. Thus, the hybrid compounds which exhibited an increase in fluorescent intensity with radical reduction can be used in the quantitative measurement of reducing species such as Fe 2+ and ascorbic acid, and hydroxyl radicals. Because these hybrid compounds have the advantage of fluorescing at longer wavelengths—661 (Cy5) or 568 (Cy3) nm, respectively, they can be used to measure radical-reducing species or radicals either in solution or in vivo.

  15. 1,3-Alternate calix[4]arene nitronyl nitroxide tetraradical and diradical: synthesis, X-ray crystallography, paramagnetic NMR spectroscopy, EPR spectroscopy, and magnetic studies

    SciTech Connect

    Rajca, Andrzej; Pink, Maren; Mukherjee, Sumit; Rajca, Suchada; Das, Kausik

    2008-04-02

    Calix[4]arenes constrained to 1,3-alternate conformation and functionalized at the upper rim with four and two nitronyl nitroxides have been synthesized, and characterized by X-ray crystallography, magnetic resonance (EPR and {sup 1}H NMR) spectroscopy, and magnetic studies. Such calix[4]arene tetraradicals and diradicals provide scaffolds for through-bond and through-space intramolecular exchange couplings.

  16. Perturbation of nuclear spin polarizations in solid state NMR of nitroxide-doped samples by magic-angle spinning without microwaves

    PubMed Central

    Thurber, Kent R.; Tycko, Robert

    2014-01-01

    We report solid state 13C and 1H nuclear magnetic resonance (NMR) experiments with magic-angle spinning (MAS) on frozen solutions containing nitroxide-based paramagnetic dopants that indicate significant perturbations of nuclear spin polarizations without microwave irradiation. At temperatures near 25 K, 1H and cross-polarized 13C NMR signals from 15N,13C-labeled L-alanine in trinitroxide-doped glycerol/water are reduced by factors as large as six compared to signals from samples without nitroxide doping. Without MAS or at temperatures near 100 K, differences between signals with and without nitroxide doping are much smaller. We attribute most of the reduction of NMR signals under MAS near 25 K to nuclear spin depolarization through the cross-effect dynamic nuclear polarization mechanism, in which three-spin flips drive nuclear polarizations toward equilibrium with spin polarization differences between electron pairs. When T1e is sufficiently long relative to the MAS rotation period, the distribution of electron spin polarization across the nitroxide electron paramagnetic resonance lineshape can be very different from the corresponding distribution in a static sample at thermal equilibrium, leading to the observed effects. We describe three-spin and 3000-spin calculations that qualitatively reproduce the experimental observations. PMID:24832263

  17. Effect of Solution Ionic Strength on the pKa of the Nitroxide pH EPR Probe 2,2,3,4,5,5-Hexamethylimidazolidin-1-oxyl.

    PubMed

    Margita, Kaleigh; Voinov, Maxim A; Smirnov, Alex I

    2017-02-17

    Spin probe and spin labeling Electron Paramagnetic Resonance methods are indispensable research tools for solving a wide range of bioanalytical problems-from measuring microviscosity and polarity of phase-separated liquids to oxygen concentrations in tissues. One of the emerging uses of spin probes are the studies of proton transfer-related and surface electrostatic phenomena. The latter Electron Paramagnetic Resonance methods rely on molecular probes containing an additional functionality capable of reversible ionization (protonation, in particular) in the immediate proximity to an Electron Paramagnetic Resonance-active reporter group, such as (N-O(•)) for nitroxides. The consequent formation of protonated and nonprotonated nitroxide species with different magnetic parameters (A iso, g iso) could be readily distinguished by Electron Paramagnetic Resonance. Bioanalytical Electron Paramagnetic Resonance studies employing pH-sensitive paramagnetic probes typically involve determination of the equilibrium constant (pK a) between the protonated and nonprotonated forms of the nitroxide. However, any chemical equilibrium involving charged species, such as ionization of acids and bases, and so the reversible protonation of the nitroxide, is known to be affected by an ionic strength of the solution. Currently, only scarce data for the effect of the solution ionic strength on the experimental pK a's of the ionizable nitroxides can be found in the literature. Here we have carried out a series of Electron Paramagnetic Resonance titration experiments for aqueous solutions of 2,2,3,4,5,5-hexamethylimidazolidin-1-oxyl (HMI) nitroxide known for one of the largest differences in the isotropic nitrogen hyperfine coupling constant A iso between the protonated and nonprotonated forms. Electrolyte concentration was varied over an exceptionally large range (i.e., from 0.05 to 5.0 M) to elucidate the effect of ionic strength on the ionization constant of this pH-sensitive Electron

  18. Distance measurements across randomly distributed nitroxide probes from the temperature dependence of the electron spin phase memory time at 240 GHz

    NASA Astrophysics Data System (ADS)

    Edwards, Devin T.; Takahashi, Susumu; Sherwin, Mark S.; Han, Songi

    2012-10-01

    At 8.5 T, the polarization of an ensemble of electron spins is essentially 100% at 2 K, and decreases to 30% at 20 K. The strong temperature dependence of the electron spin polarization between 2 and 20 K leads to the phenomenon of spin bath quenching: temporal fluctuations of the dipolar magnetic fields associated with the energy-conserving spin "flip-flop" process are quenched as the temperature of the spin bath is lowered to the point of nearly complete spin polarization. This work uses pulsed electron paramagnetic resonance (EPR) at 240 GHz to investigate the effects of spin bath quenching on the phase memory times (TM) of randomly-distributed ensembles of nitroxide molecules below 20 K at 8.5 T. For a given electron spin concentration, a characteristic, dipolar flip-flop rate (W) is extracted by fitting the temperature dependence of TM to a simple model of decoherence driven by the spin flip-flop process. In frozen solutions of 4-Amino-TEMPO, a stable nitroxide radical in a deuterated water-glass, a calibration is used to quantify average spin-spin distances as large as r¯=6.6 nm from the dipolar flip-flop rate. For longer distances, nuclear spin fluctuations, which are not frozen out, begin to dominate over the electron spin flip-flop processes, placing an effective ceiling on this method for nitroxide molecules. For a bulk solution with a three-dimensional distribution of nitroxide molecules at concentration n, we find W∝n∝1/r, which is consistent with magnetic dipolar spin interactions. Alternatively, we observe W∝n for nitroxides tethered to a quasi two-dimensional surface of large (Ø ˜ 200 nm), unilamellar, lipid vesicles, demonstrating that the quantification of spin bath quenching can also be used to discern the geometry of molecular assembly or organization.

  19. GS-Nitroxide (JP4-039)-Mediated Radioprotection of Human Fanconi Anemia Cell Lines

    PubMed Central

    Bernard, Mark E.; Kim, Hyun; Berhane, Hebist; Epperly, Michael W.; Franicola, Darcy; Zhang, Xichen; Houghton, Frank; Shields, Donna; Wang, Hong; Bakkenist, Christopher J.; Frantz, Marie-Celine; Forbeck, Erin M.; Goff, Julie P.; Wipf, Peter; Greenberger, Joel S.

    2011-01-01

    Fanconi anemia (FA) is an inherited disorder characterized by defective DNA repair and cellular sensitivity to DNA crosslinking agents. Clinically, FA is associated with high risk for marrow failure, leukemia and head and neck squamous cell carcinoma (HNSCC). Radiosensitivity in FA patients compromises the use of total-body irradiation for hematopoietic stem cell transplantation and radiation therapy for HNSCC. A radioprotector for the surrounding tissue would therefore be very valuable during radiotherapy for HNSCC. Clonogenic radiation survival curves were determined for pre- or postirradiation treatment with the parent nitroxide Tempol or JP4-039 in cells of four FA patient-derived cell lines and two transgene-corrected subclonal lines. FancG–/– (PD326) and FancD2–/– (PD20F) patient lines were more sensitive to the DNA crosslinking agent mitomycin C (MMC) than their transgene-restored subclonal cell lines (both P < 0.0001). FancD2–/– cells were more radiosensitive than the transgene restored subclonal cell line (ñ = 2.0 ± 0.7 and 4.7 ± 2.2, respectively, P = 0.03). In contrast, FancG–/– cells were radioresistant relative to the transgene-restored subclonal cell line (ñ = 9.4 ± 1.5 and 2.2 ± 05, respectively, P = 0.001). DNA strand breaks measured by the comet assay correlated with radiosensitivity. Cell lines from a Fanc-C and Fanc-A patients showed radiosensitivity similar to that of Fanc-D2–/– cells. A fluorophore-tagged JP4-039 (BODIPY-FL) analog targeted the mitochondria of the cell lines. Preirradiation or postirradiation treatment with JP4-039 at a lower concentration than Tempol significantly increased the radioresistance and stabilized the antioxidant stores of all cell lines. Tempol increased the toxicity of MMC in FancD2–/– cells. These data provide support for the potential clinical use of JP4-039 for normal tissue radioprotection during chemoradiotherapy in FA patients. PMID:21939290

  20. GS-nitroxide (JP4-039)-mediated radioprotection of human Fanconi anemia cell lines.

    PubMed

    Bernard, Mark E; Kim, Hyun; Berhane, Hebist; Epperly, Michael W; Franicola, Darcy; Zhang, Xichen; Houghton, Frank; Shields, Donna; Wang, Hong; Bakkenist, Christopher J; Frantz, Marie-Celine; Forbeck, Erin M; Goff, Julie P; Wipf, Peter; Greenberger, Joel S

    2011-11-01

    Fanconi anemia (FA) is an inherited disorder characterized by defective DNA repair and cellular sensitivity to DNA crosslinking agents. Clinically, FA is associated with high risk for marrow failure, leukemia and head and neck squamous cell carcinoma (HNSCC). Radiosensitivity in FA patients compromises the use of total-body irradiation for hematopoietic stem cell transplantation and radiation therapy for HNSCC. A radioprotector for the surrounding tissue would therefore be very valuable during radiotherapy for HNSCC. Clonogenic radiation survival curves were determined for pre- or postirradiation treatment with the parent nitroxide Tempol or JP4-039 in cells of four FA patient-derived cell lines and two transgene-corrected subclonal lines. FancG(-/-) (PD326) and FancD2(-/-) (PD20F) patient lines were more sensitive to the DNA crosslinking agent mitomycin C (MMC) than their transgene-restored subclonal cell lines (both P < 0.0001). FancD2(-/-) cells were more radiosensitive than the transgene restored subclonal cell line (ñ = 2.0 ± 0.7 and 4.7 ± 2.2, respectively, P = 0.03). In contrast, FancG(-/-) cells were radioresistant relative to the transgene-restored subclonal cell line (ñ = 9.4 ± 1.5 and 2.2 ± 05, respectively, P = 0.001). DNA strand breaks measured by the comet assay correlated with radiosensitivity. Cell lines from a Fanc-C and Fanc-A patients showed radiosensitivity similar to that of Fanc-D2(-/-) cells. A fluorophore-tagged JP4-039 (BODIPY-FL) analog targeted the mitochondria of the cell lines. Preirradiation or postirradiation treatment with JP4-039 at a lower concentration than Tempol significantly increased the radioresistance and stabilized the antioxidant stores of all cell lines. Tempol increased the toxicity of MMC in FancD2(-/-) cells. These data provide support for the potential clinical use of JP4-039 for normal tissue radioprotection during chemoradiotherapy in FA patients.

  1. Dynamics of intramolecular spin exchange interaction of a nitronyl nitroxide diradical in solution and on surfaces

    NASA Astrophysics Data System (ADS)

    Lloveras, V.; Badetti, E.; Veciana, J.; Vidal-Gancedo, J.

    2016-02-01

    In this paper we report the study of the dynamics of a thermally modulated intramolecular spin exchange interaction of a novel diradical nitronyl nitroxide-substituted disulfide in solution and when it is grafted on a gold surface. The structure of this diradical was designed to have flexible chains leading to intramolecular collisions and hence spin exchange interaction, and with an appropriate binding group to be grafted on the gold surface to study its behavior on the surface. In solution, this diradical shows a strong spin exchange interaction between both radicals which is modulated by temperature, but also gold nanoparticles (AuNPs) functionalized with this diradical permit investigation of such a phenomenon in surface-grafted radicals. The spin-labelled AuNP synthesis was optimized to obtain high coverage of spin labels to lead to high spin exchange interaction. The obtained AuNPs were studied by Electron Paramagnetic Resonance (EPR), UV-Vis, and IR spectroscopies, HR-TEM microscopy, Cyclic Voltammetry (CV), Energy Dispersive X-ray analysis (EDX) and Thermogravimetric Analysis (TGA). This inorganic-organic hybrid material also showed dipolar interactions between its radicals which were confirmed by the appearance in the EPR spectra of an |Δms| = 2 transition at half-field. This signal gives direct evidence of the presence of a high-spin state and permitted us to study the nature of the magnetic coupling between the spins which was found to be antiferromagnetic. Self-Assembled Monolayers (SAMs) of these radicals on the Au (111) substrate were also prepared and studied by contact angle, X-Ray Photoelectron Spectroscopy (XPS), Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS), Cyclic Voltammetry and EPR. The magnetic as well as the electrochemical properties of the hybrid surfaces were studied and compared with the properties of this diradical in solution. Analogies between the properties of AuNPs with high coverage of radicals and those of SAM were

  2. Dynamical transition in molecular glasses and proteins observed by spin relaxation of nitroxide spin probes and labels

    NASA Astrophysics Data System (ADS)

    Golysheva, Elena A.; Shevelev, Georgiy Yu.; Dzuba, Sergei A.

    2017-08-01

    In glassy substances and biological media, dynamical transitions are observed in neutron scattering that manifests itself as deviations of the translational mean-squared displacement, , of hydrogen atoms from harmonic dynamics. In biological media, the deviation occurs at two temperature intervals, at ˜100-150 K and at ˜170-230 K, and it is attributed to the motion of methyl groups in the former case and to the transition from harmonic to anharmonic or diffusive motions in the latter case. In this work, electron spin echo (ESE) spectroscopy—a pulsed version of electron paramagnetic resonance—is applied to study the spin relaxation of nitroxide spin probes and labels introduced in molecular glass former o-terphenyl and in protein lysozyme. The anisotropic contribution to the rate of the two-pulse ESE decay, ΔW, is induced by spin relaxation appearing because of restricted orientational stochastic molecular motion; it is proportional to <α2>τc, where <α2> is the mean-squared angle of reorientation of the nitroxide molecule around the equilibrium position and τc is the correlation time of reorientation. The ESE time window allows us to study motions with τc < 10-7 s. For glassy o-terphenyl, the <α2>τc temperature dependence shows a transition near 240 K, which is in agreement with the literature data on . For spin probes of essentially different size, the obtained data were found to be close, which evidences that motion is cooperative, involving a nanocluster of several neighboring molecules. For the dry lysozyme, the <α2>τc values below 260 K were found to linearly depend on the temperature in the same way as it was observed in neutron scattering for . As spin relaxation is influenced only by stochastic motion, the harmonic motions seen in ESE must be overdamped. In the hydrated lysozyme, ESE data show transitions near 130 K for all nitroxides, near 160 K for the probe located in the hydration layer, and near 180 K for the label in the

  3. Neuronal Cell Death and Degeneration through Increased Nitroxidative Stress and Tau Phosphorylation in HIV-1 Transgenic Rats

    PubMed Central

    Cho, Young-Eun; Lee, Myoung-Hwa; Song, Byoung-Joon

    2017-01-01

    The underlying mechanisms for increased neurodegeneration and neurocognitive deficits in HIV-infected people are unclear. Therefore, this study was aimed to investigate the mechanisms of increased neurodegeneration in 5-month old male HIV-1 Transgenic (Tg) rats compared to the age- and gender-matched wild-type (WT) by evaluating histological changes and biochemical parameters of the key proteins involved in the cell death signaling and apoptosis. Histological and immunohistochemical analyses revealed decreased neuronal cells with elevated astrogliosis in HIV-1 Tg rats compared to WT. Mechanistic studies revealed that increased levels of nitroxidative stress marker proteins such as NADPH-oxidase, cytochrome P450-2E1 (CYP2E1), inducible nitric oxide synthase (iNOS), the stress-activated mitogen-activated protein kinases such as JNK and p38K, activated cell-cycle dependent CDK5, hypoxia-inducible protein-1α, nitrated proteins, hyperphosphorylated tau, and amyloid plaques in HIV-Tg rats were consistently observed in HIV-1 Tg rats. Confocal microscopy and cell viability analyses showed that treatment with an antioxidant N-acetylcysteine or a specific inhibitor of iNOS 1400W significantly prevented the increased apoptosis of neuro-2A cells by HIV-1 Tat or gp120 protein, demonstrating the causal role of HIV-1 mediated nitroxidative stress and protein nitration in promoting neuronal cell death. Immunoprecipitation and immunoblot analysis confirmed nitration of Hsp90, evaluated as an example of nitrated proteins, suggesting possible involvement of nitrated proteins in neuronal damage. Further, activated p-JNK directly binds tau and phosphorylates multiple amino acids, suggesting an important role of p-JNK in tau hyperphosphorylation and tauopathy. These changes were accompanied with elevated levels of many apoptosis-related proteins Bax and cleaved (activated) caspase-3 as well as proinflammatory cytokines including TNF-α, IL-6 and MCP-1. Collectively, these results

  4. Formation of dense nitroxide radical layers on the Au(1 1 1) substrate for ESN-STM measurement

    NASA Astrophysics Data System (ADS)

    Krukowski, P.; Kozlowski, W.; Olejniczak, W.; Klusek, Z.; Puchalski, M.; Dabrowski, P.; Kowalczyk, P. J.; Gwozdzinski, K.; Grabowski, G.

    2008-12-01

    Ultra high vacuum scanning tunnelling microscopy and electron paramagnetic resonance spectroscopy were used to investigate ISL and TEMPOL piperidine nitroxides molecules deposited on the Au(1 1 1) substrate by the drop-cast method. The STM results suggest that both compounds form dense layers on Au(1 1 1) with high molecular mobility observed during imaging process. High resolution STM topographies of both compounds suggest a well-defined molecular order which can be ascribed to the presence of self-assembly mechanism during layers formation. The EPR results indicated that both compounds deposited on the Au(1 1 1) substrate were not reduced retaining their paramagnetic properties. The significance of the results obtained in the field of single spin detection is briefly outlined.

  5. Giant Exchange Coupling Evidenced with a Magnetization Jump at 52 T for a Gadolinium-Nitroxide Chelate.

    PubMed

    Kanetomo, Takuya; Kihara, Takumi; Miyake, Atsushi; Matsuo, Akira; Tokunaga, Masashi; Kindo, Koichi; Nojiri, Hiroyuki; Ishida, Takayuki

    2017-03-20

    The Gd-radical complex [Gd(III)(hfac)3(6bpyNO)] (6bpyNO = 2,2'-bipyridin-6-yl tert-butyl nitroxide; Hhfac = 1,1,1,5,5,5-hexafluoropentane-2,4-dione) showed a magnetization jump at 52 T observed in a pulsed-field facility, corresponding to an exchange coupling constant of -17.4 K. Furthermore, hysteretic behavior due to a relatively slow magnetization reversal was recorded around 2 T. From the high-frequency EPR study, the exchange coupling between Gd and radical spins accompanies an anisotropic character, which is responsible for both the broad jump and the slow magnetization reversal.

  6. DEER Sensitivity between Iron Centers and Nitroxides in Heme-Containing Proteins Improves Dramatically Using Broadband, High-Field EPR

    PubMed Central

    2016-01-01

    This work demonstrates the feasibility of making sensitive nanometer distance measurements between Fe(III) heme centers and nitroxide spin labels in proteins using the double electron–electron resonance (DEER) pulsed EPR technique at 94 GHz. Techniques to measure accurately long distances in many classes of heme proteins using DEER are currently strongly limited by sensitivity. In this paper we demonstrate sensitivity gains of more than 30 times compared with previous lower frequency (X-band) DEER measurements on both human neuroglobin and sperm whale myoglobin. This is achieved by taking advantage of recent instrumental advances, employing wideband excitation techniques based on composite pulses and exploiting more favorable relaxation properties of low-spin Fe(III) in high magnetic fields. This gain in sensitivity potentially allows the DEER technique to be routinely used as a sensitive probe of structure and conformation in the large number of heme and many other metalloproteins. PMID:27035368

  7. Performance of the coupled cluster and DFT methods for through-space magnetic interactions of nitroxide dimer

    NASA Astrophysics Data System (ADS)

    Saito, Toru; Ito, Akira; Watanabe, Tomohiro; Kawakami, Takashi; Okumura, Mitsutaka; Yamaguchi, Kizashi

    2012-07-01

    We have performed comparisons of several theoretical methods focusing on the through-space interactions of the F2 molecule and nitroxide (dihydronitrosyl) dimer based on the exchange coupling constant. The restricted Hartree-Fock (RHF) based coupled cluster full singles, doubles, and triples (RHF-CCSDT) and approximations to it do not show the exponential decay with increasing intermolecular distance for both antiferromagnetic and ferromagnetic interactions. On the other hand, Mukherjee's multireference CCSD (MkCCSD) method and the unrestricted HF (UHF) based CCSD (UHF-CCSD) with an approximate spin-projection (AP) method give reliable results. The present benchmark calculations are expected to contribute to the selection of an appropriate density functional theory (DFT) exchange-correlation functionals for larger systems.

  8. Cross-talk between lipid and protein carbonylation in a dynamic cardiomyocyte model of mild nitroxidative stress.

    PubMed

    Griesser, Eva; Vemula, Venukumar; Raulien, Nora; Wagner, Ulf; Reeg, Sandra; Grune, Tilman; Fedorova, Maria

    2017-04-01

    Reactive oxygen and nitrogen species (ROS/RNS) play an important role in the regulation of cardiac function. Increase in ROS/RNS concentration results in lipid and protein oxidation and is often associated with onset and/or progression of many cardiovascular disorders. However, interplay between lipid and protein modifications has not been simultaneously studied in detail so far. Biomolecule carbonylation is one of the most common biomarkers of oxidative stress. Using a dynamic model of nitroxidative stress we demonstrated rapid changes in biomolecule carbonylation in rat cardiomyocytes. Levels of carbonylated species increased as early as 15min upon treatment with the peroxynitrite donor, 3-morpholinosydnonimine (SIN-1), and decreased to values close to control after 16h. Total (lipids+proteins) vs. protein-specific carbonylation showed different dynamics, with a significant increase in protein-bound carbonyls at later time points. Treatment with SIN-1 in combination with inhibitors of proteasomal and autophagy/lysosomal degradation pathways allowed confirmation of a significant role of the proteasome in the degradation of carbonylated proteins, whereas lipid carbonylation increased in the presence of autophagy/lysosomal inhibitors. Electrophilic aldehydes and ketones formed by lipid peroxidation were identified and relatively quantified using LC-MS/MS. Molecular identity of reactive species was used for data-driven analysis of their protein targets. Combination of different enrichment strategies with LC-MS/MS analysis allowed identification of more than 167 unique proteins with 332 sites modified by electrophilic lipid peroxidation products. Gene ontology analysis of modified proteins demonstrated enrichment of several functional categories including proteins involved in cytoskeleton, extracellular matrix, ion channels and their regulation. Using calcium mobilization assays, the effect of nitroxidative stress on the activity of several ion channels was further

  9. High sensitivity and versatility of the DEER experiment on nitroxide radical pairs at Q-band frequencies.

    PubMed

    Polyhach, Yevhen; Bordignon, Enrica; Tschaggelar, René; Gandra, Sandhya; Godt, Adelheid; Jeschke, Gunnar

    2012-08-14

    Measurement of distances with the Double Electron-Electron Resonance (DEER) experiment at X-band frequencies using a pair of nitroxides as spin labels is a popular biophysical tool for studying function-related conformational dynamics of proteins. The technique is intrinsically highly precise and can potentially access the range from 1.5 to 6-10 nm. However, DEER performance drops strongly when relaxation rates of the nitroxide spin labels are high and available material quantities are low, which is usually the case for membrane proteins reconstituted into liposomes. This leads to elevated noise levels, very long measurement times, reduced precision, and a decrease of the longest accessible distances. Here we quantify the performance improvement that can be achieved at Q-band frequencies (34.5 GHz) using a high-power spectrometer. More than an order of magnitude gain in sensitivity is obtained with a homebuilt setup equipped with a 150 W TWT amplifier by using oversized samples. The broadband excitation enabled by the high power ensures that orientation selection can be suppressed in most cases, which facilitates extraction of distance distributions. By varying pulse lengths, Q-band DEER can be switched between orientationally non-selective and selective regimes. Because of suppression of nuclear modulations from matrix protons and deuterons, analysis of the Q-band data is greatly simplified, particularly in cases of very small DEER modulation depth due to low binding affinity between proteins forming a complex or low labelling efficiency. Finally, we demonstrate that a commercial Q-band spectrometer can be readily adjusted to the high-power operation.

  10. Photoinduced electron donor/acceptor processes in colloidal II-VI semiconductor quantum dots and nitroxide free radicals

    NASA Astrophysics Data System (ADS)

    Dutta, Poulami

    Electron transfer (ET) processes are one of the most researched topics for applications ranging from energy conversion to catalysis. An exciting variation is utilizing colloidal semiconductor nanostructures to explore such processes. Semiconductor quantum dots (QDs) are emerging as a novel class of light harvesting, emitting and charge-separation materials for applications such as solar energy conversion. Detailed knowledge of the quantitative dissociation of the photogenerated excitons and the interfacial charge- (electron/hole) transfer is essential for optimization of the overall efficiency of many such applications. Organic free radicals are the attractive counterparts for studying ET to/from QDs because these undergo single-electron transfer steps in reversible fashion. Nitroxides are an exciting class of stable organic free radicals, which have recently been demonstrated to be efficient as redox mediators in dye-sensitized solar cells, making them even more interesting for the aforementioned studies. This dissertation investigates the interaction between nitroxide free radicals TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl), 4-amino-TEMPO (4-amino- 2,2,6,6-tetramethylpiperidine-1-oxyl) and II-VI semiconductor (CdSe and CdTe) QDs. The nature of interaction in these hybrids has been examined through ground-state UV-Vis absorbance, steady state and time-resolved photoluminescence (PL) spectroscopy, transient absorbance, upconversion photoluminescence spectroscopy and electron paramagnetic resonance (EPR). The detailed analysis of the PL quenching indicates that the intrinsic charge transfer is ultrafast however, the overall quenching is still limited by the lower binding capacities and slower diffusion related kinetics. Careful analysis of the time resolved PL decay kinetics reveal that the decay rate constants are distributed and that the trap states are involved in the overall quenching process. The ultrafast hole transfer from CdSe QDs to 4-Amino TEMPO observed

  11. ESR lineshape and 1H spin-lattice relaxation dispersion in propylene glycol solutions of nitroxide radicals - Joint analysis

    NASA Astrophysics Data System (ADS)

    Kruk, D.; Hoffmann, S. K.; Goslar, J.; Lijewski, S.; Kubica-Misztal, A.; Korpała, A.; Oglodek, I.; Kowalewski, J.; Rössler, E. A.; Moscicki, J.

    2013-12-01

    Electron Spin Resonance (ESR) spectroscopy and Nuclear Magnetic Relaxation Dispersion (NMRD) experiments are reported for propylene glycol solutions of the nitroxide radical: 4-oxo-TEMPO-d16 containing 15N and 14N isotopes. The NMRD experiments refer to 1H spin-lattice relaxation measurements in a broad frequency range (10 kHz-20 MHz). A joint analysis of the ESR and NMRD data is performed. The ESR lineshapes give access to the nitrogen hyperfine tensor components and the rotational correlation time of the paramagnetic molecule. The NMRD data are interpreted in terms of the theory of paramagnetic relaxation enhancement in solutions of nitroxide radicals, recently presented by Kruk et al. [J. Chem. Phys. 138, 124506 (2013)]. The theory includes the effect of the electron spin relaxation on the 1H relaxation of the solvent. The 1H relaxation is caused by dipole-dipole interactions between the electron spin of the radical and the proton spins of the solvent molecules. These interactions are modulated by three dynamic processes: relative translational dynamics of the involved molecules, molecular rotation, and electron spin relaxation. The sensitivity to rotation originates from the non-central positions of the interacting spin in the molecules. The electronic relaxation is assumed to stem from the electron spin-nitrogen spin hyperfine coupling, modulated by rotation of the radical molecule. For the interpretation of the NMRD data, we use the nitrogen hyperfine coupling tensor obtained from ESR and fit the other relevant parameters. The consistency of the unified analysis of ESR and NMRD, evaluated by the agreement between the rotational correlation times obtained from ESR and NMRD, respectively, and the agreement of the translation diffusion coefficients with literature values obtained for pure propylene glycol, is demonstrated to be satisfactory.

  12. Reactive Oxygen Species and the Aging Eye: Specific Role of Metabolically Active Mitochondria in Maintaining Lens Function and in the Initiation of the Oxidation-Induced Maturity Onset Cataract--A Novel Platform of Mitochondria-Targeted Antioxidants With Broad Therapeutic Potential for Redox Regulation and Detoxification of Oxidants in Eye Diseases.

    PubMed

    Babizhayev, Mark A; Yegorov, Yegor E

    2016-01-01

    The aging eye appears to be at considerable risk from oxidative stress. A great deal of research indicates that dysfunctional mitochondria are the primary site of reactive oxygen species (ROS). More than 95% of O2 produced during normal metabolism is generated by the electron transport chain in the inner mitochondrial membrane. Mitochondria are also the major target of ROS. Cataract formation, the opacification of the eye lens, is one of the leading causes of human blindness worldwide, accounting for 47.8% of all causes of blindness. Cataracts result from the deposition of aggregated proteins in the eye lens and lens fiber cell plasma membrane damage, which causes clouding of the lens, light scattering, and obstruction of vision. ROS-induced damage in the lens cell may consist of oxidation of proteins, DNA damage, and/or lipid peroxidation, all of which have been implicated in cataractogenesis. This article is an attempt to integrate how mitochondrial ROS are altered in the aging eye along with those protective and repair therapeutic systems believed to regulate ROS levels in ocular tissues and how damage to these systems contributes to age-onset eye disease and cataract formation. Mitochondria-targeted antioxidants might be used to effectively prevent ROS-induced oxidation of lipids and proteins in the inner mitochondrial membrane in vivo. As a result of the combination of weak metal chelating, OH and lipid peroxyl radicals scavenging, reducing activities to liberated fatty acid, and phospholipid hydroperoxides, carnosine and carcinine appear to be physiological antioxidants able to efficiently protect the lipid phase of biologic membranes and aqueous environments and act as the antiapoptotic natural drug compounds The authors developed and patented the new ophthalmic compositions, including N-acetylcarnosine, acting as a prodrug of naturally targeted to mitochondria L-carnosine endowed with pluripotent antioxidant activities combined with mitochondria-targeted

  13. Extension of the AMBER force field for nitroxide radicals and combined QM/MM/PCM approach to the accurate determination of EPR parameters of DMPOH in solution

    PubMed Central

    Hermosilla, Laura; Prampolini, Giacomo; Calle, Paloma; García de la Vega, José Manuel; Brancato, Giuseppe; Barone, Vincenzo

    2015-01-01

    A computational strategy that combines both time-dependent and time-independent approaches is exploited to accurately model molecular dynamics and solvent effects on the isotropic hyperfine coupling constants of the DMPO-H nitroxide. Our recent general force field for nitroxides derived from AMBER ff99SB is further extended to systems involving hydrogen atoms in β-positions with respect to NO. The resulting force-field has been employed in a series of classical molecular dynamics simulations, comparing the computed EPR parameters from selected molecular configurations to the corresponding experimental data in different solvents. The effect of vibrational averaging on the spectroscopic parameters is also taken into account, by second order vibrational perturbation theory involving semi-diagonal third energy derivatives together first and second property derivatives. PMID:26584116

  14. Evaluation of spin labels for in-cell EPR by analysis of nitroxide reduction in cell extract of Xenopus laevis oocytes

    NASA Astrophysics Data System (ADS)

    Azarkh, Mykhailo; Okle, Oliver; Eyring, Philipp; Dietrich, Daniel R.; Drescher, Malte

    2011-10-01

    Spin-label electron paramagnetic resonance (SL-EPR) spectroscopy has become a powerful and useful tool for studying structure and dynamics of biomacromolecules. However, utilizing these methods at physiological temperatures for in-cell studies is hampered by reduction of the nitroxide spin labels and thus short half-lives in the cellular environment. Consequently, reduction kinetics of two structurally different nitroxides was investigated in cell extracts of Xenopus laevis oocytes using rapid-scan cw-experiments at X-band. The five member heterocyclic ring nitroxide PCA (3-carboxy-2,2,5,5-tetramethylpyrrolidinyl-1-oxy) under investigation features much higher stability against intracellular reduction than the six member ring analog TOAC (2,2,6,6-tetramethylpiperidine-N-oxyl-4-amino-4-carboxilic acid) and is therefore a suitable spin label type for in-cell EPR. The kinetic data can be described according to the Michaelis-Menten model and thus suggest an enzymatic or enzyme-mediated reduction process.

  15. New insights into the mechanism of amine/nitroxide cycling during the hindered amine light stabilizer inhibited oxidative degradation of polymers.

    PubMed

    Gryn'ova, Ganna; Ingold, K U; Coote, Michelle L

    2012-08-08

    High-level ab initio molecular orbital theory calculations are used to identify the origin of the remarkably high inhibition stoichiometric factors exhibited by dialkylamine-based radical-trapping antioxidants. We have calculated the free energy barriers and reaction energies at 25, 80, and 260 °C in the gas phase and in aqueous solution for a broad range of reactions that might, potentially, be involved in amine/nitroxide cycling, as well as several novel pathways proposed as part of the present work, including that of N-alkyl hindered amine light stabilizer activation. We find that most of the literature nitroxide regeneration cycles should be discarded on either kinetic or thermodynamic grounds; some are even inconsistent with existing experimental observations. We therefore propose a new mechanistic cycle that relies on abstraction of a β-hydrogen atom from an alkoxyamine (R(1)R(2)NOCHR(3)R(4)). Our results suggest that this cycle is energetically feasible for a range of substrates and provides an explanation for previously misinterpreted or unexplained experimental results. We also explore alternative mechanisms for amine/nitroxide cycling for cases where the alkoxyamines do not possess an abstractable β-hydrogen.

  16. Cytochrome P450-2E1 is involved in aging-related kidney damage in mice through increased nitroxidative stress.

    PubMed

    Abdelmegeed, Mohamed A; Choi, Youngshim; Ha, Seung-Kwoon; Song, Byoung-Joon

    2017-08-24

    The aim of this study was to investigate the role of cytochrome P450-2E1 (CYP2E1) in aging-dependent kidney damage since it is poorly understood. Young (7 weeks) and aged female (16-17 months old) wild-type (WT) and Cyp2e1-null mice were used. Kidney histology showed that aged WT mice exhibited typical signs of kidney aging such as cell vacuolation, inflammatory cell infiltration, cellular apoptosis, glomerulonephropathy, and fibrosis, along with significantly elevated levels of renal TNF-α and serum creatinine than all other groups. Furthermore, the highest levels of renal hydrogen peroxide, protein carbonylation and nitration were observed in aged WT mice. These increases in the aged WT mice were accompanied by increased levels of iNOS and mitochondrial nitroxidative stress through altered amounts and activities of the mitochondrial complex proteins and significantly reduced levels of the antioxidant glutathione (GSH). In contrast, the aged Cyp2e1-null mice exhibited significantly higher antioxidant capacity with elevated heme oxygenase-1 and catalase activities compared to all other groups, while maintaining normal GSH levels with significantly less mitochondrial nitroxidative stress compared to the aged WT mice. Thus, CYP2E1 is important in causing aging-related kidney damage most likely through increasing nitroxidative stress and that CYP2E1 could be a potential target in preventing aging-related kidney diseases. Published by Elsevier Ltd.

  17. Endothelial NO and O2•− Production Rates Differentially Regulate Oxidative, Nitroxidative and Nitrosative Stress in the Microcirculation

    PubMed Central

    Kar, Saptarshi; Kavdia, Mahendra

    2013-01-01

    Endothelial dysfunction causes an imbalance in endothelial NO and O2•− production rates and increased peroxynitrite formation. Peroxynitrite and its decomposition products cause multiple deleterious effects including tyrosine nitration of proteins, superoxide dismutase (SOD) inactivation and tissue damage. Studies have shown that peroxynitrite formation during endothelial dysfunction is strongly dependent on the NO and O2•− production rates. Previous experimental and modeling studies examining the role of NO and O2•− production imbalance on peroxynitrite formation showed different results in biological and synthetic systems. However, there is a lack of quantitative information about the formation and biological relevance of peroxynitrite in oxidative, nitroxidative and nitrosative stress conditions in the microcirculation. We developed a computational biotransport model to examine the role of endothelium NO and O2•− production on the complex biochemical NO and O2•− interactions in the microcirculation. We also modeled the effect of variability in SOD expression and activity during oxidative stress. The results showed that peroxynitrite concentration increased with increase in either O2•− to NO or NO to O2•− production rate ratios (QO2•−/QNO or QNO/QO2•−, respectively). The peroxynitrite concentrations were similar for both production rate ratios indicating that peroxynitrite related nitroxidative and nitrosative stresses may be similar in endothelial dysfunction or iNOS induced NO production. The endothelial peroxynitrite concentration increased with increase in both QO2•−/QNO or QNO/QO2•− ratios at SOD concentrations 0.1–100 μM. The absence of SOD may not mitigate the extent of peroxynitrite mediated toxicity as we predicted insignificant increase in peroxynitrite levels beyond QO2•−/QNO and QNO/QO2•−ratio of 1. The results support the experimental observations of biological systems and show that peroxynitrite

  18. Magneto-Structural Relationship on Strong Exchange Interactions between Chelating Nitroxide Radical and Transition-Metal Spins

    NASA Astrophysics Data System (ADS)

    Okazawa, A.

    2017-05-01

    To develop room-temperature molecule-based magnets, metal-radical approach is one of the most promising strategies. Strong magnetic interactions beyond a thermal energy of room temperature are essential for achieving such coveted materials. We have developed 2-pyridyl tert-butyl nitroxide (2pyNO) derivatives as a chelating radical ligand with a copper(II) or nickel(II) ion. The torsion angle (ϕ) around M-O-N-C2py is a useful metric in describing correlation between the structure and metal-radical exchange coupling 2J (defined as H = -2JΣSi ·Si +1). Although we previously reported such a magneto-structural correlation, in this paper we updated the plot of 2J versus ϕ by dividing into two categories: the hexacoordinatenickel(II) and copper(II) complexes. An empirical linear correlation was formulated for both Cu and Ni categories, as 2J/k B = a + b|ϕ| with a = 503(112) K and b = -36(3) K deg-1 and with a = 449(46) K and b = -21.0(17) K deg-1, respectively. In the latter plot we confirm that the relationship can be generalized for the oxoverdazyl and dithiadiazolyl systems as well as the 2pyNO systems. The results are supported by the calculations based on the density functional theory.

  19. Observation of steric hindrance effect controlling crystal packing structures and physical properties in three new isomeric nitronyl nitroxide radicals

    NASA Astrophysics Data System (ADS)

    Zhao, Hai-Rong; Sun, Jia-Sen; Sui, Yun-Xia; Ren, Xiao-Ming; Yao, Bin-Qian; Shen, Lin-Jiang; Meng, Qing-Jin

    2009-07-01

    Three isomeric nitronyl nitroxide radical compounds, 2-[ n-( N-benzyl)pyridinium]-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide bromide ( n = 2, 3 and 4 for 1, 2 and 3, respectively), have been synthesized and structurally characterized. The influence of steric hindrance on the molecular packing structures and physical properties has been observed. In the radical 1, such steric hindrance leads to a folding conformation of the imidazoline and benzene rings and the intramolecular C-H…π interaction between the methyl group and the benzene ring. There is no such effect in 2 and 3. In crystal of 2, there are the intermolecular C-H…π between methyl groups and benzene ring and intermolecular π…π stacking interaction between pyridine and benzene rings. Crystal of 2 with a chiral space group P2 12 12 1 shows the SHG response about 0.4 times as that of urea. In crystal of 3, there are three symmetry-independent radical molecules, which form an unusually six-membered supramolecular ring via intermolecular O…π interactions. For the solid sample of 3, the X-band EPR exhibits an axially symmetric signal and magnetic susceptibility data suggest intermolecular antiferromagnetic (AFM) coupling interactions and very weak intermolecular ferromagnetic (FM) coupling interactions which is more likely caused by magnetic anisotropy, while measurements of both 1 and 2 show isotropic X-band EPR signals and simple Currie-Weiss magnetic behavior.

  20. Small Molecule GS-Nitroxide Ameliorates Ionizing Irradiation-Induced Delay in Bone Wound Healing in a Novel Murine Model

    PubMed Central

    Gokhale, Abhay; Rwigema, Jean-Claude; Epperly, Michael; Glowacki, Julie; Wang, Hong; Wipf, Peter; Goff, Julie P.; Dixon, Tracy; Patrene, Ken; Greenberger, Joel S.

    2010-01-01

    We studied radioprotection and mitigation by mitochondrial-targeted Tempol (GS-nitroxide, JP4-039), in a mouse injury/irradiation model of combined injury (fracture/irradiation). Right hind legs of control C57BL/6NHsd female mice, mice pretreated with MnSOD-PL, JP4-039, or with amifostine were irradiated with single and fractionated doses of 0 to 20 Gy. Twenty-four hours later, unicortical holes were drilled into the tibiae of both hind legs; at intervals, tibias were excised, radiographed, and processed for histology. Bone wounds irradiated to 20 or 30 Gy showed delayed healing at 21 to 28 days. Treatment with JP4-039 MnSOD-PL or amifostine, before or after single fraction 20 Gy or during fractionated irradiation followed by drilling accelerated wound healing at days 21 and 28. Orthotopic 3LL tumors were not protected by JP4-039 or amifostine. In nonirradiated mice, pretreatment with JP4-039 accelerated bone wound healing. This test system should be useful for the development of new small molecule radioprotectors. PMID:20668303

  1. Molecular dynamics and partitioning of di-tert-butyl nitroxide in stratum corneum membranes: effect of terpenes.

    PubMed

    Camargos, Heverton Silva; Silva, Adolfo Henrique Moraes; Anjos, Jorge Luiz Vieira; Alonso, Antonio

    2010-05-01

    In this work, we have used electron paramagnetic resonance (EPR) spectroscopy of the small spin label di-tert-butyl nitroxide (DTBN), which partitions the aqueous and hydrocarbon phases, to study the interaction of the terpenes alpha-terpineol, 1,8-cineole, L(-)-carvone and (+)-limonene with the uppermost skin layer, the stratum corneum, and the membrane models of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC). The EPR spectra indicated that the terpenes increase both the partition coefficient and the rotational correlation time of the spin labels in the stratum corneum membranes, whereas similar effects were observed in the DMPC and DPPC bilayers only at temperatures below the liquid-crystalline phase. The EPR parameter associated to probe polarity inside the membranes showed thermotropically induced changes, suggesting relocations of spin probe, which were dependent on the membrane phases. While the DMPC and DPPC bilayers showed abrupt changes in the partitioning and rotational correlation time parameters in the phase transitions, the SC membranes were characterized by slight changes in the total range of measured temperatures, presenting the greatest changes or membranes reorganizations in the temperature range of approximately 50 to approximately 74 degrees C. The results suggest that terpenes act as spacers, weakening the hydrogen-bonded network at the polar interface and thus fluidizing the stratum corneum lipids.

  2. Homoleptic Ce(III) and Ce(IV) Nitroxide Complexes: Significant Stabilization of the 4+ Oxidation State

    SciTech Connect

    Bogart, Justin A.; Lewis, Andrew J.; Medling, Scott A.; Piro, Nicholas A.; Carroll, Patrick J.; Booth, Corwin H.; Schelter, Eric J.

    2014-06-25

    Electrochemical experiments performed on the complex Ce-IV[2-((BuNO)-Bu-t)py](4), where [2-((BuNO)-Bu-t)py](-) = N-tert-butyl-N-2-pyridylnitroxide, indicate a 2.51 V stabilization of the 4+ oxidation state of Ce compared to [(Bu4N)-Bu-n](2)[Ce(NO3)(6)] in acetonitrile and a 2.95 V stabilization compared to the standard potential for the ion under aqueous conditions. Density functional theory calculations suggest that this preference for the higher oxidation state is a result of the tetrakis(nitroxide) ligand framework at the Ce cation, which allows for effective electron donation into, and partial covalent overlap with, vacant 4f orbitals with delta symmetry. The results speak to the behavior of CeO2 and related solid solutions in oxygen uptake and transport applications, in particular an inherent local character of bonding that stabilizes the 4+ oxidation state. The results indicate a cerium(IV) complex that has been stabilized to an unprecedented degree through tuning of its ligand-field environment.

  3. Small molecule GS-nitroxide ameliorates ionizing irradiation-induced delay in bone wound healing in a novel murine model.

    PubMed

    Gokhale, Abhay; Rwigema, Jean-Claude; Epperly, Michael W; Glowacki, Julie; Wang, Hong; Wipf, Peter; Goff, Julie P; Dixon, Tracy; Patrene, Ken; Greenberger, Joel S

    2010-01-01

    We studied radioprotection and mitigation by mitochondrial-targeted Tempol (GS-nitroxide, JP4-039), in a mouse injury/irradiation model of combined injury (fracture/irradiation). Right hind legs of control C57BL/6NHsd female mice, mice pretreated with MnSOD-PL, JP4-039, or with amifostine were irradiated with single and fractionated doses of 0 to 20 Gy. Twenty-four hours later, unicortical holes were drilled into the tibiae of both hind legs; at intervals, tibias were excised, radiographed, and processed for histology. Bone wounds irradiated to 20 or 30 Gy showed delayed healing at 21 to 28 days. Treatment with JP4-039 MnSOD-PL or amifostine, before or after single fraction 20 Gy or during fractionated irradiation followed by drilling accelerated wound healing at days 21 and 28. Orthotopic 3LL tumors were not protected by JP4-039 or amifostine. In nonirradiated mice, pretreatment with JP4-039 accelerated bone wound healing. This test system should be useful for the development of new small molecule radioprotectors.

  4. New spectral-spatial imaging algorithm for full EPR spectra of multiline nitroxides and pH sensitive trityl radicals

    NASA Astrophysics Data System (ADS)

    Tseitlin, Mark; Biller, Joshua R.; Elajaili, Hanan; Khramtsov, Valery V.; Dhimitruka, Ilirian; Eaton, Gareth R.; Eaton, Sandra S.

    2014-08-01

    An algorithm is derived and demonstrated that reconstructs an EPR spectral-spatial image from projections with arbitrarily selected gradients. This approach permits imaging wide spectra without the use of the very large sweep widths and gradients that would be required for spectral-spatial imaging with filtered back projection reconstruction. Each projection is defined as the sum of contributions at the set of locations in the object. At each location gradients shift the spectra in the magnetic field domain, which is equivalent to a phase change in the Fourier-conjugate frequency domain. This permits solution of the problem in the frequency domain. The method was demonstrated for 2D images of phantoms consisting of (i) two tubes containing 14N and 15N nitroxide and (ii) two tubes containing a pH sensitive trityl radical at pH 7.0 and 7.2. In each case spectral slices through the image agree well with the full spectra obtained in the absence of gradient.

  5. Computation of DNP coupling factors of a nitroxide radical in toluene: seamless combination of MD simulations and analytical calculations.

    PubMed

    Sezer, Deniz

    2013-01-14

    Dynamic nuclear polarization (DNP) employs paramagnetic species to increase the NMR signal of nuclear spins. In liquids, the efficiency of the effect depends on the strength of the interaction between the electron and nuclear spins and the time scales on which this interaction is modulated by the physical motion of the spin-bearing molecules. An approach to quantitatively predict the contribution of molecular motions to the DNP enhancement using molecular dynamics (MD) simulations is developed and illustrated for the nitroxide radical TEMPOL in liquid toluene. A multi-resolution strategy that combines explicit treatment of the solvent at short distances from the free radical with implicit description at large intermolecular distances is adopted. Novel analytical expressions are obtained to correct for the finite spatial extent of the MD simulations. The atomistic and analytical descriptions are sewn seamlessly together by ensuring that for molecular trajectories that start in the near (explicit) region and end in the distant (implicit) region the analytical dipolar spectral densities reproduce the MD estimates. The spectral densities obtained from the developed approach are used to calculate DNP coupling factors separately for the ring and methyl protons of toluene. The agreement with previously reported experimental DNP data at a magnetic field of 3.4 T is noteworthy and encouraging. Maximum obtainable DNP enhancements at other magnetic fields are predicted.

  6. Rationale for mitochondria-targeting strategies in cancer bioenergetic therapies.

    PubMed

    Jose, Caroline; Rossignol, Rodrigue

    2013-01-01

    In the 1920s, Otto Warburg first hypothesized that mitochondrial impairment is a leading cause of cancer although he recognized the existence of oxidative tumors. Likewise, Weinhouse and others in the 50s found that deficient mitochondrial respiration is not an obligatory feature of cancer and Peter Vaupel suggested in the 1990s that tumor oxygenation rather than OXPHOS capacity was the limiting factor of mitochondrial energy production in cancer. Recent studies now clearly indicate that mitochondria are highly functional in mice tumors and the field of oncobioenergetic identified MYC, Oct1 and RAS as pro-OXPHOS oncogenes. In addition, cancer cells adaptation to aglycemia, metabolic symbiosis between hypoxic and non-hypoxic tumor regions as well the reverse Warburg hypothesis support the crucial role of mitochondria in the survival of a subclass of tumors. Therefore, mitochondria are now considered as potential targets for anti-cancer therapy and tentative strategies including a bioenergetic profile characterization of the tumor and the subsequent adapted bioenergetic modulation could be considered for cancer killing. This article is part of a Directed Issue entitled: Bioenergetic dysfunction, adaptation and therapy.

  7. Neuroprotection and Anti-Epileptogenesis with Mitochondria-Targeted Antioxidant

    DTIC Science & Technology

    2015-10-01

    tetanus toxin (Tx) model of epilepsy . Progress on the project has been limited throughout the grant period due to an inability to obtain a sufficient...of epilepsy . The pilocarpine-induced model of status epilepticus (PILO) will be used to test SS-31 as a neuroprotectant, the kindling model will be... epilepsy . The combination of the negative results we obtained with SS-31 treatment in the PILO and kindling models and our continued difficulty in

  8. Neuroprotection and Anti-Epileptogenesis with Mitochondria-Targeted Antioxidant

    DTIC Science & Technology

    2014-10-01

    efficacy of a novel, mitochondrial-targeted antioxidant SS-31, as a neuroprotective and antiepileptogenic agent in three experimental models of epilepsy ...anticonvulsant properties of SS-31 in the Tetanus Toxin (TX) model of mesial temporal lobe epilepsy . The TX model is established and run on a regular basis in

  9. Antimutagenic activity of mitochondria-targeted plastoquinone derivative.

    PubMed

    Chistyakov, V A; Sazykina, M A; Alexandrova, A A; Belichenko, N I; Mashkina, E V; Gutnikova, L V; Zolotukhin, P V; Shkurat, T P

    2010-03-01

    The ability of cationic plastoquinone derivative 10-(6'-plastoquinonyl) decyltriphenylphosphonium (SkQ1) to modify processes of spontaneous and induced mutagenesis was studied. It is shown that daily introduction of this compound into male Wistar rats in doses of 25 and 250 nmol/kg during two weeks decreases spontaneous level of chromosome aberrations in anaphase in the eye cornea from 0.39 +/- 0.09 to 0.13 +/- 0.08 and 0.14 +/- 0.05, respectively. The level of 8-hydroxy-2'-deoxyguanosine in blood serum of the investigated animals decreases from 32.12 +/- 1.55 to 25.90 +/- 2.26 and 25.76 +/- 1.50 ng/ml, respectively. These facts indicate that the decrease in spontaneous clastogenesis is caused by decreased level of DNA damage by endogenous reactive oxygen species. A higher dose of SkQ1 also decreases to control level chromosome aberrations caused by oxygen under pressure of 0.5 MPa for 60 min. It is also shown in experiments with bacterial biosensors that SkQ1 is able to efficiently protect cells against genotoxic effect of UV radiation at 300-400 nm.

  10. Comparing continuous wave progressive saturation EPR and time domain saturation recovery EPR over the entire motional range of nitroxide spin labels.

    PubMed

    Nielsen, Robert D; Canaan, Stephane; Gladden, James A; Gelb, Michael H; Mailer, Colin; Robinson, Bruce H

    2004-07-01

    The measurement of spin-lattice relaxation rates from spin labels, such as nitroxides, in the presence and absence of spin relaxants provides information that is useful for determining biomolecular properties such as nucleic acid dynamics and the interaction of proteins with membranes. We compare X-band continuous wave (CW) and pulsed or time domain (TD) EPR methods for obtaining spin-lattice relaxation rates of spin labels across the entire range of rotational motion to which relaxation rates are sensitive. Model nitroxides and spin-labeled biological species are used to illustrate the potential complications that arise in extracting relaxation data under conditions typical to biological experiments. The effect of super hyperfine (SHF) structure is investigated for both CW and TD spectra. First and second harmonic absorption and dispersion CW spectra of the nitroxide spin label, TEMPOL, are all fit simultaneously to a model of SHF structure over a range of microwave amplitudes. The CW spectra are novel because all harmonics and microwave phases were acquired simultaneously using our homebuilt CW/TD spectrometer. The effect of the SHF structure on the pulsed free induction decay (FID) and pulsed saturation recovery spectrum is shown for both protonated and deuterated TEMPOL. We present novel pulsed saturation recovery measurements on biological molecules, including spin-lattice relaxation rates of spin-labeled proteins and spin-labeled double-stranded DNA. The impact of structure and dynamics on relaxation rates are discussed in the context of each of these examples. Collisional relaxation rates with oxygen and transition metal paramagnetic relaxants are extracted using both continuous wave and time domain methods. The extent of the errors inherent in the CW method and the advantages of pulsed methods for unambiguously measuring collisional relaxation rates are discussed. Spin-lattice relaxation rates, determined by both CW and pulsed methods, are used to determine

  11. Comparing continuous wave progressive saturation EPR and time domain saturation recovery EPR over the entire motional range of nitroxide spin labels

    NASA Astrophysics Data System (ADS)

    Nielsen, Robert D.; Canaan, Stephane; Gladden, James A.; Gelb, Michael H.; Mailer, Colin; Robinson, Bruce H.

    2004-07-01

    The measurement of spin-lattice relaxation rates from spin labels, such as nitroxides, in the presence and absence of spin relaxants provides information that is useful for determining biomolecular properties such as nucleic acid dynamics and the interaction of proteins with membranes. We compare X-band continuous wave (CW) and pulsed or time domain (TD) EPR methods for obtaining spin-lattice relaxation rates of spin labels across the entire range of rotational motion to which relaxation rates are sensitive. Model nitroxides and spin-labeled biological species are used to illustrate the potential complications that arise in extracting relaxation data under conditions typical to biological experiments. The effect of super hyperfine (SHF) structure is investigated for both CW and TD spectra. First and second harmonic absorption and dispersion CW spectra of the nitroxide spin label, TEMPOL, are all fit simultaneously to a model of SHF structure over a range of microwave amplitudes. The CW spectra are novel because all harmonics and microwave phases were acquired simultaneously using our homebuilt CW/TD spectrometer. The effect of the SHF structure on the pulsed free induction decay (FID) and pulsed saturation recovery spectrum is shown for both protonated and deuterated TEMPOL. We present novel pulsed saturation recovery measurements on biological molecules, including spin-lattice relaxation rates of spin-labeled proteins and spin-labeled double-stranded DNA. The impact of structure and dynamics on relaxation rates are discussed in the context of each of these examples. Collisional relaxation rates with oxygen and transition metal paramagnetic relaxants are extracted using both continuous wave and time domain methods. The extent of the errors inherent in the CW method and the advantages of pulsed methods for unambiguously measuring collisional relaxation rates are discussed. Spin-lattice relaxation rates, determined by both CW and pulsed methods, are used to determine

  12. Single-molecule magnetic behavior in a neutral terbium(III) complex of a picolinate-based nitronyl nitroxide free radical.

    PubMed

    Coronado, Eugenio; Giménez-Saiz, Carlos; Recuenco, Alejandro; Tarazón, Ana; Romero, Francisco M; Camón, Agustín; Luis, Fernando

    2011-08-15

    The terdentate anionic picolinate-based nitronyl nitroxide (picNN) free radical forms neutral and robust homoleptic complexes with rare earth-metal ions. The nonacoordinated Tb(3+) complex Tb(picNN)(3)·6H(2)O is a single-molecule magnet with an activation energy barrier Δ = 22.8 ± 0.5 K and preexponential factor τ(0) = (5.5 ± 1.1) × 10(-9) s. It shows magnetic hysteresis below 1 K. © 2011 American Chemical Society

  13. Three-dimensionally ordered macroporous nitroxide polymer brush electrodes prepared by surface-initiated atom transfer polymerization for organic radical batteries.

    PubMed

    Lin, Chun-Hao; Chou, Wei-Jen; Lee, Jyh-Tsung

    2012-01-01

    The synthesis and electrochemical performance of three-dimensionally ordered macroporous (3DOM) nitroxide polymer brush electrodes for organic radical batteries are reported. The 3DOM electrodes are synthesized via polystyrene colloidal crystal templating with electropolymerization of polypyrrole, modification of surface initiator, and surface-initiated atom transfer radical polymerization. The discharge capacity of the 3DOM electrodes is proportional to the thickness of the inverse opal. The discharge capacity of the 3DOM electrode at a discharge rate of 5 C is 40 times higher than that of the planar electrode; its cycle-life performance exhibits 96.1% retention after 250 cycles.

  14. Solution Dynamics of Redox Noninnocent Nitrosoarene Ligands: Mapping the Electronic Criteria for the Formation of Persistent Metal-Coordinated Nitroxide Radicals.

    PubMed

    Barnett, Brandon R; Labios, Liezel A; Moore, Curtis E; England, Jason; Rheingold, Arnold L; Wieghardt, Karl; Figueroa, Joshua S

    2015-07-20

    The redox-noninnocence of metal-coordinated C-organo nitrosoarenes has been established on the basis of solid-state characterization techniques, but the solution-phase properties of this class of metal-coordinated radicals have been relatively underexplored. In this report, the solution-phase properties and dynamics of the bis-nitrosobenzene diradical complex trans-Pd(κ(1)-N-PhNO)2(CNAr(Dipp2))2 are presented. This complex, which is best described as containing singly reduced phenylnitroxide radical ligands, is shown to undergo facile nitrosobenzene dissociation in solution to form the metalloxaziridine Pd(η(2)-N,O-PhNO)(CNAr(Dipp2))2 and thus is not a persistent species in solution. An equilibrium between trans-Pd(κ(1)-N-PhNO)2(CNAr(Dipp2))2, Pd(η(2)-N,O-PhNO)(CNAr(Dipp2))2, and free nitrosobenzene is established in solution, with the metalloxaziridine being predominantly favored. Efforts to perturb this equilibrium by the addition of excess nitrosobenzene reveal that the formation of trans-Pd(κ(1)-N-PhNO)2(CNAr(Dipp2))2 is in competition with insertion-type chemistry of Pd(η(2)-N,O-PhNO)(CNAr(Dipp2))2 and is therefore not a viable strategy for the production of a kinetically persistent bis-nitroxide radical complex. Electronic modification of the nitrosoarene framework was explored as a means to generate a persistent trans-Pd(κ(1)-N-ArNO)2(CNAr(Dipp2))2 complex. While most substitution schemes failed to significantly perturb the kinetic lability of the nitrosoarene ligands in the corresponding trans-Pd(κ(1)-N-ArNO)2(CNAr(Dipp2))2 complexes, utilization of para-formyl or para-cyano nitrosobenzene produced bis-nitroxide diradical complexes that display kinetic persistence in solution. The origin of this persistence is rationalized by the ability of para-formyl- and para-cyano-aryl groups to both attenuate the trans effect of the corresponding nitrosoarene and, more importantly, delocalize spin density away from the aryl-nitroxide NO unit. The results

  15. Smooth transition between SMM and SCM-type slow relaxing dynamics for a 1-D assemblage of {Dy(nitronyl nitroxide)2} units.

    PubMed

    Liu, Ruina; Li, Licun; Wang, Xiaoling; Yang, Peipei; Wang, Chao; Liao, Daizheng; Sutter, Jean-Pascal

    2010-04-21

    A model example for size effects on the dynamic susceptibility behavior is provided by the chain compound [{Dy(hfac)(3)NitPhIm(2)}Dy(hfac)(3)] (NitPhIm = 2-[4-(1-imidazole)phenyl]nitronyl nitroxide radical). The Arrhenius plot reveals two relaxation regimes attributed to SMM (Delta = 17.1 K and tau(0) = 17.5 x 10(-6) s) and SCM (Delta = 82.7 K and tau(0) = 8.8 x 10(-8) s) behaviors. The ferromagnetic exchange among the spin carriers has been established for the corresponding Gd derivative.

  16. Mitochondrial-Targeted Nitroxides Disrupt Mitochondrial Architecture and Inhibit Expression of Peroxiredoxin 3 and FOXM1 in Malignant Mesothelioma Cells

    PubMed Central

    Cunniff, Brian; Benson, Kira; Stumpff, Jason; Newick, Kheng; Held, Paul; Taatjes, Douglas; Joseph, Joy; Kalyanaraman, Balaraman; Heintz, Nicholas H.

    2013-01-01

    Malignant mesothelioma (MM) is an intractable tumor of the peritoneal and pleural cavities primarily linked to exposure to asbestos. Recently, we described an interplay between mitochondrial-derived oxidants and expression of FOXM1, a redox-responsive transcription factor that has emerged as a promising therapeutic target in solid malignancies. Here we have investigated the effects of nitroxides targeted to mitochondria via triphenylphosphonium (TPP) moieties on mitochondrial oxidant production, expression of FOXM1 and peroxiredoxin 3 (PRX3), and cell viability in MM cells in culture. Both Mito-carboxy-proxyl (MCP) and Mito-TEMPOL (MT) caused dose-dependent increases in mitochondrial oxidant production that was accompanied by inhibition of expression of FOXM1 and PRX3 and loss of cell viability. At equivalent concentrations TPP, CP, and TEMPOL had no effect on these endpoints. Live cell ratiometric imaging with a redox-responsive green fluorescent protein targeted to mitochondria (mito-roGFP) showed that MCP and MT, but not CP, TEMPOL, or TPP, rapidly induced mitochondrial fragmentation and swelling, morphological transitions that were associated with diminished ATP levels and increased production of mitochondrial oxidants. Mdivi-1, an inhibitor of mitochondrial fission, did not rescue mitochondria from fragmentation by MCP. Immunofluorescence microscopy experiments indicate a fraction of FOXM1 coexists in the cytoplasm with mitochondrial PRX3. Our results indicate that MCP and MT inhibit FOXM1 expression and MM tumor cell viability via perturbations in redox homeostasis caused by marked disruption of mitochondrial architecture, and suggest that both compounds, either alone or in combination with thiostrepton or other agents, may provide credible therapeutic options for the management of MM. PMID:23018647

  17. An advanced approach to the evaluation of the spin-rotational term for a nitronyl nitroxide in fluid solution

    NASA Astrophysics Data System (ADS)

    Collauto, A.; Barbon, A.; Zerbetto, M.; Brustolon, M.

    2013-10-01

    In this work we focus on the spin-rotational contribution to transverse relaxation rate 1/T2 for a nitronyl nitroxide radical (PTIO) in fluid solution of toluene. We recorded the X-band continuous-wave electron paramagnetic resonance spectra of the radical in a wide temperature range and compared them with the calculated spectra. The calculation was afforded in two steps: first, we calculated the spectral profiles in the same range of temperatures taking into account the hyperfine interaction with the two 14N nuclei, by integration of the stochastic Liouville equation with the E-SpiReS package in the presence of the tumbling motion of the molecule, which modulates the hyperfine and the g tensors. In the second step, we included the proton hyperfine structure by convoluting the spectrum with the pattern due to the 17 PTIO-coupled protons. A further Lorentzian broadening was added by a best fitting procedure to reproduce the experimental linewidths. The additional broadening is associated with a relaxation rate, Wγ, which varies linearly on kB?; this trend is expected for the spin-rotational relaxation term modelled by the well-known Atkins-Kivelson expression. We found a discrepancy between the two parameters associated with the radii of the radical, obtained either from the rotational diffusion tensor in the framework of the Debye-Stokes-Einstein model or from the spin-rotational contribution. We discuss this issue in relation to the intrinsic approximations of the spin-rotational model and, in particular, the isotropic Brownian rotation.

  18. Solid-phase supported profluorescent nitroxide probe for the determination of aerosol-borne reactive oxygen species.

    PubMed

    Sleiman, Mohamad; Destaillats, Hugo; Gundel, Lara A

    2013-11-15

    Reactive oxygen species (ROS) and free radicals play important roles in the chemical transformation and adverse health effects of environmental aerosols. This work presents a simple and sensitive method for sampling and analysis of ROS using a packed column coated with a profluorescent nitroxide scavenger, proxyl fluorescamine (PF). Quantification was performed by extraction and analysis using HPLC with fluorescence detection. For comparison, the conventional method of collecting aerosols into dichlorofluorescin (DCFH) aqueous solution was used as a reference. The method was successfully applied to the determination of ROS in a model secondary organic aerosol (SOA) system generated by ozonolysis of nicotine, as well as in secondhand tobacco smoke (SHS). ROS concentrations between 50-565 nmol m(-3) were detected in fresh SOA and SHS samples. After SHS aging for 22 h, 13-18% of the initial ROS mass remained, suggesting the presence of persistent ROS. The new method offers better stability and reproducibility along with sensitivity comparable to that of DCFH (method detection limit of 3.2 and 1.4 nmol m(-3) of equivalent H2O2 for PF and DCFH respectively). The PF probe was stable during storage at room temperature and not reactive with ozone or NOx, whereas DCFH in the particle-collecting liquid system was strongly influenced by ozone and NOx interferences. This case study provides a good basis for employing solid-phase supported PF for field measurement of specific ROS in other combustion systems (i.e. biomass burning, candles, and diesel exhaust) and environmental aerosols. © 2013 Elsevier B.V. All rights reserved.

  19. EPR Studies of Functionally Active, Nitroxide Spin-Labeled Peptide Analogs of the C-terminus of a G-Protein Alpha Subunit

    PubMed Central

    Van Eps, Ned; Anderson, Lori L.; Kisselev, Oleg G.; Baranski, Thomas J.; Hubbell, Wayne L.; Marshall, Garland R.

    2010-01-01

    The C-terminal tail of the transducin alpha subunit, Gtα(340–350), is known to bind and stabilize the active conformation of rhodopsin upon photoactivation (R*). Five spin-labeled analogs of Gtα(340–350) demonstrated native-like activity in their ability to bind and stabilize R*. The spin label 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid (TOAC) was employed at interior sites within the peptide, whereas a Proxyl (3-carboxyl-2,2,5,5-tetramethyl-pyrrolidinyloxy) spin label was employed at the amino terminus of the peptide. Upon binding to R*, the electron paramagnetic resonance spectrum of TOAC343-Gtα(340–350) revealed greater immobilization of the nitroxide when compared to that of the N-terminal modified Proxyl-Gtα(340–350) analog. A double-labeled Proxyl/TOAC348-Gtα(340–350) was examined by DEER spectroscopy to determine the distribution of distances between the two nitroxides in the peptides when in solution and when bound to R*. TOAC and Proxyl spin labels in this GPCR-G-protein α-peptide system provide unique biophysical probes that can be used to explore the structure and conformational changes at the rhodopsin-G-protein interface. PMID:20695526

  20. The application of profluorescent nitroxides to detect reactive oxygen species derived from combustion-generated particulate matter: Cigarette smoke - A case study

    NASA Astrophysics Data System (ADS)

    Miljevic, B.; Fairfull-Smith, K. E.; Bottle, S. E.; Ristovski, Z. D.

    2010-06-01

    Reactive oxygen species (ROS) and related free radicals are considered to be key factors underpinning the various adverse health effects associated with exposure to ambient particulate matter. Therefore, measurement of ROS is a crucial factor for assessing the potential toxicity of particles. In this work, a novel profluorescent nitroxide, BPEAnit, was investigated as a probe for detecting particle-derived ROS. BPEAnit has a very low fluorescence emission due to inherent quenching by the nitroxide group, but upon radical trapping or redox activity, a strong fluorescence is observed. BPEAnit was tested for detection of ROS present in mainstream and sidestream cigarette smoke. In the case of mainstream cigarette smoke, there was a linear increase in fluorescence intensity with an increasing number of cigarette puffs, equivalent to an average of 101 nmol ROS per cigarette based on the number of moles of the probe reacted. Sidestream cigarette smoke sampled from an environmental chamber exposed BPEAnit to much lower concentrations of particles, but still resulted in a clearly detectible increase in fluorescence intensity with sampling time. It was calculated that the amount of ROS was equivalent to 50 ± 2 nmol per mg of particulate matter; however, this value decreased with ageing of the particles in the chamber. Overall, BPEAnit was shown to provide a sensitive response related to the oxidative capacity of the particulate matter. These findings present a good basis for employing the new BPEAnit probe for the investigation of particle-related ROS generated from cigarette smoke as well as from other combustion sources.

  1. Terbium(III) and yttrium(III) complexes with pyridine-substituted nitronyl nitroxide radical and different β-diketonate ligands. Crystal structures and magnetic and luminescence properties.

    PubMed

    Lannes, Anthony; Intissar, Mourad; Suffren, Yan; Reber, Christian; Luneau, Dominique

    2014-09-15

    A terbium(III) complex of nitronyl nitroxide free radical 2-(2-pyridyl)-4,4,5,5-tetramethyl-4,5-dihydro1H-imidazolyl-1-oxy-3-oxide (NIT2Py), [Tb(acac)3NIT2Py]·0.5H2O (3) (acac = acetylacetonate), was synthesized for comparison with the previously reported [Tb(hfac)3NIT2Py]·0.5C7H16 (1) (hfac = hexafluoroacetylacetonate), together with their yttrium analogues [Y(hfac)3NIT2Py]·0.5C7H16 (2) and [Y(acac)3NIT2Py]·0.5H2O (4). The crystal structures show that in all complexes the nitronyl nitroxide radical acts as a chelating ligand. Magnetic studies show that 3 like 1 exhibits slow relaxation of magnetization at low temperature, suggesting single-molecule magnet behavior. The luminescence spectra show resolved vibronic structure with the main interval decreasing from 1600 cm(-1) to 1400 cm(-1) between 80 and 300 K. This effect is analyzed quantitatively using experimental Raman frequencies.

  2. Ultra-short laser-accelerated proton pulses have similar DNA-damaging effectiveness but produce less immediate nitroxidative stress than conventional proton beams

    PubMed Central

    Raschke, S.; Spickermann, S.; Toncian, T.; Swantusch, M.; Boeker, J.; Giesen, U.; Iliakis, G.; Willi, O.; Boege, F.

    2016-01-01

    Ultra-short proton pulses originating from laser-plasma accelerators can provide instantaneous dose rates at least 107-fold in excess of conventional, continuous proton beams. The impact of such extremely high proton dose rates on A549 human lung cancer cells was compared with conventionally accelerated protons and 90 keV X-rays. Between 0.2 and 2 Gy, the yield of DNA double strand breaks (foci of phosphorylated histone H2AX) was not significantly different between the two proton sources or proton irradiation and X-rays. Protein nitroxidation after 1 h judged by 3-nitrotyrosine generation was 2.5 and 5-fold higher in response to conventionally accelerated protons compared to laser-driven protons and X-rays, respectively. This difference was significant (p < 0.01) between 0.25 and 1 Gy. In conclusion, ultra-short proton pulses originating from laser-plasma accelerators have a similar DNA damaging potential as conventional proton beams, while inducing less immediate nitroxidative stress, which probably entails a distinct therapeutic potential. PMID:27578260

  3. Molecular Dynamics Simulation Study of Solvent and State of Charge Effects on Solid-Phase Structure and Counterion Binding in a Nitroxide Radical Containing Polymer Energy Storage Material

    SciTech Connect

    Kemper, Travis W.; Gennett, Thomas; Larsen, Ross E.

    2016-10-19

    Here we performed molecular dynamics simulations to understand the effects of solvent swelling and state of charge (SOC) on the redox active, organic radical cathode material poly(2,2,6,6-tetramethylpiperidinyloxy methacrylate) (PTMA). We show that the polar solvent acetonitrile primarily solvates the nitroxide radical without disrupting the packing of the (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) pendant groups of PTMA. We also simulated bulk PTMA in different SOC, 25%, 50%, 75%, and 100%, by converting the appropriate number of TEMPO groups to the cation charge state and adding BF4- counterions to the simulation. At each SOC the packing of PTMA, the solvent, and the counterions were examined. The binding of the anion to the nitroxide cation site was examined using the potential of mean force and found to be on the order of tens of meV, with a binding energy that decreased with increasing SOC. Additionally, we found that the cation state is stabilized by the presence of a nearby anion by more than 1 eV, and the implications of this stabilization on charge transport are discussed. Finally, we describe the implications of our results for how the SOC of an organic electrode affects electron and anion charge transport during the charging and discharging processes.

  4. Molecular Dynamics Simulation Study of Solvent and State of Charge Effects on Solid-Phase Structure and Counterion Binding in a Nitroxide Radical Containing Polymer Energy Storage Material

    DOE PAGES

    Kemper, Travis W.; Gennett, Thomas; Larsen, Ross E.

    2016-10-19

    Here we performed molecular dynamics simulations to understand the effects of solvent swelling and state of charge (SOC) on the redox active, organic radical cathode material poly(2,2,6,6-tetramethylpiperidinyloxy methacrylate) (PTMA). We show that the polar solvent acetonitrile primarily solvates the nitroxide radical without disrupting the packing of the (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) pendant groups of PTMA. We also simulated bulk PTMA in different SOC, 25%, 50%, 75%, and 100%, by converting the appropriate number of TEMPO groups to the cation charge state and adding BF4- counterions to the simulation. At each SOC the packing of PTMA, the solvent, and the counterions were examined.more » The binding of the anion to the nitroxide cation site was examined using the potential of mean force and found to be on the order of tens of meV, with a binding energy that decreased with increasing SOC. Additionally, we found that the cation state is stabilized by the presence of a nearby anion by more than 1 eV, and the implications of this stabilization on charge transport are discussed. Finally, we describe the implications of our results for how the SOC of an organic electrode affects electron and anion charge transport during the charging and discharging processes.« less

  5. Spatial distribution of stabilizer-derived nitroxide radicals during thermal degradation of poly(acrylonitrile-butadiene-styrene) copolymers: a unified picture from pulsed ELDOR and ESR imaging.

    PubMed

    Jeschke, Gunnar; Schlick, Shulamith

    2006-09-21

    Double Electron-Electron Resonance (DEER) provides information on the spatial distribution of radicals on the length scale of a few nanometres, while Electron Spin Resonance Imaging (ESRI) provides information on a length scale of millimetres with a resolution of about 100 micrometres. Despite the gap between these length scales, results from the two techniques are found to complement and support each other in the characterization of the identity and distribution of nitroxide radicals derived from the Hindered Amine Stabilizer (HAS) Tinuvin 770 in poly(acrylonitrile-butadiene-styrene) (ABS) copolymers. DEER measurements demonstrate that there is no significant formation of biradicals from the bifunctional HAS, and provide the distributions of local radical concentrations. These distributions are poorly resolved for model-free analysis of the DEER data by the Tikhonov regularization; the resolution was significantly improved by utilizing information obtained by ESRI. DEER data can be fitted with only one adjustable parameter, namely the average radical concentration, if 1D and 2D spectral--spatial ESRI results on both the spatial distribution of nitroxides and their distribution between the acrylonitrile--styrene-rich (SAN) and butadiene-rich (B) microphases are considered.

  6. The structure and electronic state of photoexcited fullerene linked with a nitroxide radical based on an analysis of a two-dimensional electron paramagnetic resonance nutation spectrum

    NASA Astrophysics Data System (ADS)

    Mizuochi, Norikazu; Ohba, Yasunori; Yamauchi, Seigo

    1999-08-01

    An electron paramagnetic resonance (EPR) study of 3,4-fulleropyrrolidine-2-spiro-4'[2',2',6',6'-tetramethyl]piperidine-1'-oxyl (1) was performed on the photoexcited quartet state in toluene glass. The spectrum of the |S,Ms>=|3/2,±3/2>⇔|3/2,±1/2> transitions was observed selectively by using a two-dimensional (2D) nutation method and analyzed with a spectral simulation in a randomly oriented system. A position of the nitroxide moiety was determined with respect to the zero-field splitting (zfs) axes of excited triplet fullerene (3C60) by taking into account of the dipolar-dipolar interaction between the radical and 3C60, the hyperfine coupling, the anisotropic g-value of the nitroxide radical, and the zfs of the 3C60 moiety. It was found that none of the zfs axes of the 3C60 moiety coincide with the local C2 axis of the molecule which is defined by the position of addition. A symmetry of the electronic structure in 3C60 is discussed on the basis of the result.

  7. Ultra-short laser-accelerated proton pulses have similar DNA-damaging effectiveness but produce less immediate nitroxidative stress than conventional proton beams

    NASA Astrophysics Data System (ADS)

    Raschke, S.; Spickermann, S.; Toncian, T.; Swantusch, M.; Boeker, J.; Giesen, U.; Iliakis, G.; Willi, O.; Boege, F.

    2016-08-01

    Ultra-short proton pulses originating from laser-plasma accelerators can provide instantaneous dose rates at least 107-fold in excess of conventional, continuous proton beams. The impact of such extremely high proton dose rates on A549 human lung cancer cells was compared with conventionally accelerated protons and 90 keV X-rays. Between 0.2 and 2 Gy, the yield of DNA double strand breaks (foci of phosphorylated histone H2AX) was not significantly different between the two proton sources or proton irradiation and X-rays. Protein nitroxidation after 1 h judged by 3-nitrotyrosine generation was 2.5 and 5-fold higher in response to conventionally accelerated protons compared to laser-driven protons and X-rays, respectively. This difference was significant (p < 0.01) between 0.25 and 1 Gy. In conclusion, ultra-short proton pulses originating from laser-plasma accelerators have a similar DNA damaging potential as conventional proton beams, while inducing less immediate nitroxidative stress, which probably entails a distinct therapeutic potential.

  8. Structural determinants of nitroxide motion in spin-labeled proteins: Solvent-exposed sites in helix B of T4 lysozyme

    PubMed Central

    Guo, Zhefeng; Cascio, Duilio; Hideg, Kálmán; Hubbell, Wayne L.

    2008-01-01

    Site-directed spin labeling provides a means for exploring structure and dynamics in proteins. To interpret the complex EPR spectra that often arise, it is necessary to characterize the rotamers of the spin-labeled side chain and the interactions they make with the local environment in proteins of known structure. For this purpose, crystal structures have been determined for T4 lysozyme bearing a nitroxide side chain (R1) at the solvent-exposed helical sites 41 and 44 in the B helix. These sites are of particular interest in that the corresponding EPR spectra reveal two dynamic states of R1, one of which is relatively immobilized suggesting interactions of the nitroxide with the environment. The crystal structures together with the effect of mutagenesis of nearest neighbors on the motion of R1 suggest intrahelical interactions of 41R1 with the i + 4 residue and of 44R1 with the i + 1 residue. Such interactions appear to be specific to particular rotamers of the R1 side chain. PMID:18096642

  9. Successful MALDI-MS analysis of synthetic polymers with labile end-groups: the case of nitroxide-mediated polymerization using the MAMA-SG1 alkoxyamine.

    PubMed

    Barrère, Caroline; Chendo, Christophe; Phan, Trang N T; Monnier, Valérie; Trimaille, Thomas; Humbel, Stéphane; Viel, Stéphane; Gigmes, Didier; Charles, Laurence

    2012-06-18

    A sample pretreatment was evaluated to enable the production of intact cationic species of synthetic polymers holding a labile end-group using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. More specifically, polymers obtained by nitroxide-mediated polymerization involving the MAMA-SG1 alkoxyamine were stirred for a few hours in trifluoroacetic acid (TFA) to induce the substitution of a tert-butyl group on the nitrogen of nitroxide end-group by a hydrogen atom. Nuclear magnetic resonance, electrospray ionization tandem mass spectrometry, and theoretical calculations were combined to scrutinize this sample pretreatment from both mechanistic and energetic points of view. The substitution reaction was found to increase the dissociation energy of the fragile C-ON bond to a sufficient extent to prevent this bond to be spontaneously cleaved during MALDI analysis. This TFA treatment is shown to be very efficient regardless of the nature of the polymer, as evidenced by reliable MALDI mass spectrometric data obtained for poly(ethylene oxide), polystyrene and poly(butylacrylate). Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Observation of ferromagnetic exchange, spin crossover, reductively induced oxidation, and field-induced slow magnetic relaxation in monomeric cobalt nitroxides.

    PubMed

    Gass, Ian A; Tewary, Subrata; Nafady, Ayman; Chilton, Nicholas F; Gartshore, Christopher J; Asadi, Mousa; Lupton, David W; Moubaraki, Boujemaa; Bond, Alan M; Boas, John F; Guo, Si-Xuan; Rajaraman, Gopalan; Murray, Keith S

    2013-07-01

    The reaction of [Co(II)(NO3)2]·6H2O with the nitroxide radical, 4-dimethyl-2,2-di(2-pyridyl) oxazolidine-N-oxide (L(•)), produces the mononuclear transition-metal complex [Co(II)(L(•))2](NO3)2 (1), which has been investigated using temperature-dependent magnetic susceptibility, electron paramagnetic resonance (EPR) spectroscopy, electrochemistry, density functional theory (DFT) calculations, and variable-temperature X-ray structure analysis. Magnetic susceptibility measurements and X-ray diffraction (XRD) analysis reveal a central low-spin octahedral Co(2+) ion with both ligands in the neutral radical form (L(•)) forming a linear L(•)···Co(II)···L(•) arrangement. This shows a host of interesting magnetic properties including strong cobalt-radical and radical-radical intramolecular ferromagnetic interactions stabilizing a S = (3)/2 ground state, a thermally induced spin crossover transition above 200 K and field-induced slow magnetic relaxation. This is supported by variable-temperature EPR spectra, which suggest that 1 has a positive D value and nonzero E values, suggesting the possibility of a field-induced transverse anisotropy barrier. DFT calculations support the parallel alignment of the two radical π*NO orbitals with a small orbital overlap leading to radical-radical ferromagnetic interactions while the cobalt-radical interaction is computed to be strong and ferromagnetic. In the high-spin (HS) case, the DFT calculations predict a weak antiferromagnetic cobalt-radical interaction, whereas the radical-radical interaction is computed to be large and ferromagnetic. The monocationic complex [Co(III)(L(-))2](BPh4) (2) is formed by a rare, reductively induced oxidation of the Co center and has been fully characterized by X-ray structure analysis and magnetic measurements revealing a diamagnetic ground state. Electrochemical studies on 1 and 2 revealed common Co-redox intermediates and the proposed mechanism is compared and contrasted with that of

  11. Synthesis, structure, and magnetic properties of (6-9)-nuclear Ni(II) trimethylacetates and their heterospin complexes with nitroxides.

    PubMed

    Ovcharenko, Victor; Fursova, Elena; Romanenko, Galina; Eremenko, Igor; Tretyakov, Evgeny; Ikorskii, Vladimir

    2006-07-10

    New polynuclear nickel trimethylacetates [Ni6(OH)4(C5H9O2)8(C5H10O2)4] (6), [Ni7(OH)7(C5H9O2)7(C5H10O2)6(H2O)] x 0.5 C6H14 x 0.5 H2O (7), [Ni8(OH)4(H2O)2(C5H9O2)12] (8), and [Ni9(OH)6(C5H9O2)12(C5H10O2)4] x C5H10O2 x 3 H2O (9), where C5H9O2 is trimethylacetate and C5H10O2 is trimethylacetic acid, have been found. Their structures were determined by X-ray crystallography. Because of their high solubility in low-polarity organic solvents, compounds 6-9 reacted with stable organic radicals to form the first heterospin compounds based on polynuclear Ni(II) trimethylacetate and nitronyl nitroxides containing pyrazole (L(1)-L(3)), methyl (L(4)), or imidazole (L(5)) substituent groups, respectively, in side chain [Ni7(OH)5(C5H9O2)9(C5H10O2)2(L(1))2(H2O)] x 0.5 C6H14 x H2O (6+1a), [Ni7(OH)5(C5H9O2)9(C5H10O2)2(L2)2(H2O)] x H2O (6+1b), [Ni7(OH)5(C5H9O2)9(C5H10O2)2(L(3))2(H2O)] x H2O (6+1c), [Ni6(OH)3(C5H9O2)9(C5H10O2)4(L(4))] x 1.5 C6H14 (6''), and [Ni4OH)3(C5H9O2)5(C5H10O2)4(L(5))] x 1.5 C7H8 (4). Their structures were also determined by X-ray crystallography. Although Ni(II) trimethylacetates may have varying nuclearity and can change their nuclearity during recrystallization or interactions with nitroxides, this family of compounds is easy to study because of its topological relationship. For any of these complexes, the polynuclear framework may be derived from the [Ni6] polynuclear fragment {Ni6(mu4-OH)2(mu3-OH)2(mu2-C5H9O2-O,O')6(mu2-C5H9O2-O,O)(mu4-C5H9O2-O,O,O',O')(C5H10O2)4}, which is shaped like an open book. On the basis of this fragment, the structure of 7-nuclear compounds (7 and 6+1a-c) is conveniently represented as the result of symmetric addition of other mononuclear fragments to the four Ni(II) ions lying at the vertexes of the [Ni6] open book. The 9-nuclear complex is formed by the addition of trinuclear fragments to two Ni(II) ions lying on one of the lateral edges of the [Ni6] open book. This wing of the 9-nuclear complex preserves its structure in

  12. Nitroxide derivatives of non-steroidal anti-inflammatory drugs exert anti-inflammatory and superoxide dismutase scavenging properties in A459 cells

    PubMed Central

    Flores-Santana, Wilmarie; Moody, Terry; Chen, Weibin; Gorczynski, Michael J; Shoman, Mai E; Velázquez, Carlos; Thetford, Angela; Mitchell, James B; Cherukuri, Murali K; King, S Bruce; Wink, David A

    2012-01-01

    BACKGROUND AND PURPOSE Inflammation and reactive oxygen species are associated with the promotion of various cancers. The use of non-steroidal anti-inflammatory drugs (NSAIDs) in cancer prevention treatments has been promising in numerous cancers. We report the evaluation of NSAIDs chemically modified by the addition of a redox-active nitroxide group. TEMPO-aspirin (TEMPO-ASA) and TEMPO-indomethacin (TEMPO-IND) were synthesized and evaluated in the lung cancer cell line A549. EXPERIMENTAL APPROACHES We evaluated physico-chemical properties of TEMPO-ASA and TEMPO-IND by electron paramagnetic resonance and cyclic voltammetry. Superoxide dismutase-like properties was assayed by measuring cytochrome c reduction and anti-inflammatory effects were assayed by measuring production of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4). MTT proliferation assay and clonogenic assay were evaluated in the A549 lung carcinoma cell line. Maximum tolerated doses (MTD) and acute ulcerogenic index were also evaluated in in vivo. KEY RESULTS MTD were: TEMPO (140 mg·kg−1), ASA (100 mg·kg−1), indomethacin (5 mg·kg−1), TEMPO-ASA (100 mg·kg−1) and TEMPO-IND (40 mg·kg−1). While TEMPO-ASA was as well tolerated as ASA, TEMPO-IND showed an eightfold improvement over indomethacin. TEMPO-IND showed markedly less gastric toxicity than the parent NSAID. Both TEMPO-ASA and TEMPO-IND inhibited production of PGE2 and LTB4 in A549 cells with maximum effects at 100 µg·mL−1 or 10 µg·mL−1 respectively. CONCLUSIONS AND IMPLICATIONS The nitroxide-NSAIDs retained superoxide scavenging capacity of the parent nitroxide and anti-inflammatory effects, inhibiting cyclooxygenase and 5-lipoxygenase enzymes. These redox-modified NSAIDs might be potential drug candidates, as they exhibit the pharmacological properties of the parent NSAID with antioxidant activity decreasing NSAID-associated toxicity. PMID:21658022

  13. 2,2,6,6-Tetramethylpiperydinyl-1-oxyl (TEMPO) Functionalized Benzoxazines Prepared with a One-Pot Synthesis for Reactive/Crosslinkable Initiators of Nitroxide Mediated Polymerization.

    PubMed

    Han, Yi-Jen; Liu, Ying-Ling

    2017-08-01

    In this work, the incorporation of a 2,2,6,6-tetramethylpiperydinyl-1-oxyl (TEMPO) group to a benzoxazine ring is performed using a one-pot synthesis for the preparation of TEMPO-functionalized benzoxazine compounds and polymers as reactive and crosslinkable initiators for nitroxide-mediated polymerization (NMP). The TEMPO-functionalization reaction of benzoxazine, traced with (1) H NMR, is conducted with sequential radical transfer and coupling reactions. Moreover, polystyrene-grafted polybenzoxazine copolymers are prepared with the TEMPO-benzoxazine initiator and NMP of styrene. The polymerization system exhibits the characteristics of controlled radical polymerization, including controlled molecular weights of products and ability for sequential polymerization. Moreover, based on the chemical reactivity and crosslinking ability of benzoxazine groups, the synthesis route developed in this work will widen the scope of the design and synthesis of functional and high-performance polymers. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Insights into peptide-membrane interactions of newly synthesized, nitroxide-containing analogs of the peptaibiotic trichogin GA IV using EPR.

    PubMed

    Bortolus, Marco; Dalzini, Annalisa; Maniero, Anna Lisa; Panighel, Giacomo; Siano, Alvaro; Toniolo, Claudio; De Zotti, Marta; Formaggio, Fernando

    2017-01-01

    Trichogin GA IV is a short-length (10-amino acid long), mostly hydrophobic, peptaibiotic with an N-terminal fatty acyl chain and a C-terminal 1,2-amino alcohol. A cardinal role of the terminal moieties in the cytotoxic activity of trichogin has been recently found. Previously, peptide orientation and dynamics of trichogin analogs in the membrane were studied using methyl ester derivatives. Therefore, in the present work we synthesized several trichogin analogs with naturally occurring terminal groups to verify whether these moieties have any effect on peptide-membrane interaction. These trichogin analogs, both neutral and carrying a positively charged Lys residue, bear the nitroxide-containing α-amino acid TOAC to study them using EPR spectroscopy. Vesicles were used to investigate orientation and penetration depth of the peptide at room temperature. Bicelles were employed to evaluate the order, dynamics, and orientation of the peptide at a near physiological temperature. In addition, the position of the N-terminal 1-octanoyl chain in the membrane was studied by labeling it with a nitroxide. The secondary structure of the peptides in vesicles was studied by CD spectroscopy showing that they adopt a mostly α-helical structure. In vesicles, the analogs insert below the lipid headgroups with the helix axis oriented parallel to the membrane surface at a peptide-to-lipid (P:L) ratio of 1:100. The presence of the single, positively charged Lys residue does not alter the orientation adopted by the peptides. In bicelles at P:L ratios 1:100 and 1:60, the peptide adopts a transmembrane orientation characterized by a very low orientational order, whereas at a 1:15 P:L ratio it severely disrupts the membrane. Our data shows that overall orientation and insertion in model membranes of the native trichogin GA IV are strictly comparable to those of its methyl ester analogs previously examined. © 2016 Wiley Periodicals, Inc.

  15. The internal dynamics of mini c TAR DNA probed by electron paramagnetic resonance of nitroxide spin-labels at the lower stem, the loop, and the bulge.

    PubMed

    Sun, Yan; Zhang, Ziwei; Grigoryants, Vladimir M; Myers, William K; Liu, Fei; Earle, Keith A; Freed, Jack H; Scholes, Charles P

    2012-10-30

    Electron paramagnetic resonance (EPR) at 236.6 and 9.5 GHz probed the tumbling of nitroxide spin probes in the lower stem, in the upper loop, and near the bulge of mini c TAR DNA. High-frequency 236.6 GHz EPR, not previously applied to spin-labeled oligonucleotides, was notably sensitive to fast, anisotropic, hindered local rotational motion of the spin probe, occurring approximately about the NO nitroxide axis. Labels attached to the 2'-aminocytidine sugar in the mini c TAR DNA showed such anisotropic motion, which was faster in the lower stem, a region previously thought to be partially melted. More flexible labels attached to phosphorothioates at the end of the lower stem tumbled isotropically in mini c TAR DNA, mini TAR RNA, and ψ(3) RNA, but at 5 °C, the motion became more anisotropic for the labeled RNAs, implying more order within the RNA lower stems. As observed by 9.5 GHz EPR, the slowing of nanosecond motions of large segments of the oligonucleotide was enhanced by increasing the ratio of the nucleocapsid protein NCp7 to mini c TAR DNA from 0 to 2. The slowing was most significant at labels in the loop and near the bulge. At a 4:1 ratio of NCp7 to mini c TAR DNA, all labels reported tumbling times of >5 ns, indicating a condensation of NCp7 and TAR DNA. At the 4:1 ratio, pulse dipolar EPR spectroscopy of bilabels attached near the 3' and 5' termini showed evidence of an NCp7-induced increase in the 3'-5' end-to-end distance distribution and a partially melted stem.

  16. Nitroxide spin labels as EPR reporters of the relaxation and magnetic properties of the heme-copper site in cytochrome bo3, E. coli.

    PubMed

    Oganesyan, Vasily S; White, Gaye F; Field, Sarah; Marritt, Sophie; Gennis, Robert B; Yap, Lai Lai; Thomson, Andrew J

    2010-11-01

    A nitroxide spin label (SL) has been used to probe the electron spin relaxation times and the magnetic states of the oxygen-binding heme-copper dinuclear site in Escherichia coli cytochrome bo(3), a quinol oxidase (QO), in different oxidation states. The spin lattice relaxation times, T(1), of the SL are enhanced by the paramagnetic metal sites in QO and hence show a strong dependence on the oxidation state of the latter. A new, general form of equations and a computer simulation program have been developed for the calculation of relaxation enhancement by an arbitrary fast relaxing spin system of S ≥ 1/2. This has allowed us to obtain an accurate estimate of the transverse relaxation time, T (2), of the dinuclear coupled pair Fe(III)-Cu(B)(II) in the oxidized form of QO that is too short to measure directly. In the case of the F' state, the relaxation properties of the heme-copper center have been shown to be consistent with a ferryl [Fe(IV)=O] heme and Cu(B)(II) coupled by approximately 1.5-3 cm(-1) to a radical. The magnitude suggests that the coupling arises from a radical form of the covalently linked tyrosine-histidine ligand to Cu(II) with unpaired spin density primarily on the tyrosine component. This work demonstrates that nitroxide SLs are potentially valuable tools to probe both the relaxation and the magnetic properties of multinuclear high-spin paramagnetic active sites in proteins that are otherwise not accessible from direct EPR measurements.

  17. ESR lineshape and {sup 1}H spin-lattice relaxation dispersion in propylene glycol solutions of nitroxide radicals – Joint analysis

    SciTech Connect

    Kruk, D.; Hoffmann, S. K.; Goslar, J.; Lijewski, S.; Kubica-Misztal, A.; Korpała, A.; Oglodek, I.; Moscicki, J.; Kowalewski, J.; Rössler, E. A.

    2013-12-28

    Electron Spin Resonance (ESR) spectroscopy and Nuclear Magnetic Relaxation Dispersion (NMRD) experiments are reported for propylene glycol solutions of the nitroxide radical: 4-oxo-TEMPO-d{sub 16} containing {sup 15}N and {sup 14}N isotopes. The NMRD experiments refer to {sup 1}H spin-lattice relaxation measurements in a broad frequency range (10 kHz–20 MHz). A joint analysis of the ESR and NMRD data is performed. The ESR lineshapes give access to the nitrogen hyperfine tensor components and the rotational correlation time of the paramagnetic molecule. The NMRD data are interpreted in terms of the theory of paramagnetic relaxation enhancement in solutions of nitroxide radicals, recently presented by Kruk et al. [J. Chem. Phys. 138, 124506 (2013)]. The theory includes the effect of the electron spin relaxation on the {sup 1}H relaxation of the solvent. The {sup 1}H relaxation is caused by dipole-dipole interactions between the electron spin of the radical and the proton spins of the solvent molecules. These interactions are modulated by three dynamic processes: relative translational dynamics of the involved molecules, molecular rotation, and electron spin relaxation. The sensitivity to rotation originates from the non-central positions of the interacting spin in the molecules. The electronic relaxation is assumed to stem from the electron spin–nitrogen spin hyperfine coupling, modulated by rotation of the radical molecule. For the interpretation of the NMRD data, we use the nitrogen hyperfine coupling tensor obtained from ESR and fit the other relevant parameters. The consistency of the unified analysis of ESR and NMRD, evaluated by the agreement between the rotational correlation times obtained from ESR and NMRD, respectively, and the agreement of the translation diffusion coefficients with literature values obtained for pure propylene glycol, is demonstrated to be satisfactory.

  18. Frequency Dependence of Electron Spin Relaxation Times in Aqueous Solution for a Nitronyl Nitroxide Radical and Per-deuterated-Tempone between 250 MHz and 34 GHz

    PubMed Central

    Biller, Joshua R.; Meyer, Virginia M.; Elajaili, Hanan; Rosen, Gerald M.; Eaton, Sandra S.; Eaton, Gareth R.

    2012-01-01

    Electron spin relaxation times of perdeuterated tempone (PDT) 1 and of a nitronyl nitroxide 2-(4-carboxy-phenyl)-4,4,5,5-tetramethylimidazoline-3-oxide-1-oxyl) 2 in aqueous solution at room temperature were measured by 2-pulse electron spin echo (T2) or 3-pulse inversion recovery (T1) in the frequency range of 250 MHz to 34 GHz. At 9 GHz values of T1 measured by long-pulse saturation recovery were in good agreement with values determined by inversion recovery. Below 9 GHz for 1 and below 1.5 GHz for 2, T1~ T2, as expected in the fast tumbling regime. At higher frequencies T2 was shorter than T1 due to incomplete motional averaging of g and A anisotropy. The frequency dependence of 1/T1 is modeled as the sum of spin rotation, modulation of g and A-anisotropy, and a thermally-activated process that has maximum contribution at about 1.5 GHz. The spin lattice relaxation times for the nitronyl nitroxide were longer than for PDT by a factor of about 2 at 34 GHz, decreasing to about a factor of 1.5 at 250 MHz. The rotational correlation times, τR are calculated to be 9 ps for 1 and about 25 ps for 2. The longer spin lattice relaxation times for 2 than for 1 at 9 and 34 GHz are due predominantly to smaller contributions from spin rotation that arise from slower tumbling. The smaller nitrogen hyperfine couplings for the nitronyl 2 than for 1 decrease the contribution to relaxation due to modulation of A anisotropy. However, at lower frequencies the slower tumbling of 2 results in a larger value of ωτR (ω is the resonance frequency) and larger values of the spectral density function, which enhances the contribution from modulation of anisotropic interactions for 2 to a greater extent than for 1. PMID:23123770

  19. The Internal Dynamics of Mini c TAR DNA Probed by EPR of Nitroxide Spin Labels at the Lower Stem, the Loop, and the Bulge †

    PubMed Central

    Sun, Yan; Zhang, Ziwei; Grigoryants, Vladimir M.; Myers, William K.; Liu, Fei; Earle, Keith A.; Freed, Jack H.; Scholes, Charles P.

    2012-01-01

    Electron paramagnetic resonance (EPR) at 236.6 GHz and 9.5 GHz probed the tumbling of nitroxide spin probes in the lower stem, the upper loop, and near the bulge of mini c TAR DNA. High frequency 236.6 GHz EPR, not previously applied to spin labeled oligonucleotides, was notably sensitive to fast, anisotropic, hindered local rotational motion of the spin probe, occurring approximately about the NO nitroxide axis. Labels attached to the 2′-amino cytidine sugar in the mini c TAR DNA showed such anisotropic motion, which was faster in the lower stem, a region previously suggested to be partially melted. More flexible labels attached to phosphorothioates at the end of the lower stem tumbled isotropically in mini c TAR DNA, mini TAR RNA, and ψ3 RNA, but at 5 °C the motion became more anisotropic for the labeled RNAs, implying more order within the RNA lower stems. As observed by 9.5 GHz EPR, the slowing of nanosecond motions of large segments of the oligonucleotide was enhanced by increasing the ratio of the nucleocapsid protein NCp7 to mini c TAR DNA from zero to two. The slowing was most significant at labels in the loop and near the bulge. At a 4:1 ratio of NCp7 to mini c TAR DNA all labels reported tumbling times > 5 ns, indicating a condensation of NCp7 and TAR DNA. At the 4:1 ratio, pulse dipolar EPR spectroscopy of bi-labels attached near the 3′ and 5′ terminals showed evidence for an NCp7-induced increase in the 3′ - 5 ′end-to-end distance distribution and a partially melted stem. PMID:23009298

  20. Core-shell hybrid upconversion nanoparticles carrying stable nitroxide radicals as potential multifunctional nanoprobes for upconversion luminescence and magnetic resonance dual-modality imaging

    NASA Astrophysics Data System (ADS)

    Chen, Chuan; Kang, Ning; Xu, Ting; Wang, Dong; Ren, Lei; Guo, Xiangqun

    2015-03-01

    Nitroxide radicals, such as 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO) and its derivatives, have recently been used as contrast agents for magnetic resonance imaging (MRI) and electron paramagnetic resonance imaging (EPRI). However, their rapid one-electron bioreduction to diamagnetic N-hydroxy species when administered intravenously has limited their use in in vivo applications. In this article, a new approach of silica coating for carrying stable radicals was proposed. A 4-carboxyl-TEMPO nitroxide radical was covalently linked with 3-aminopropyl-trimethoxysilane to produce a silanizing TEMPO radical. Utilizing a facile reaction based on the copolymerization of silanizing TEMPO radicals with tetraethyl orthosilicate in reverse microemulsion, a TEMPO radicals doped SiO2 nanostructure was synthesized and coated on the surface of NaYF4:Yb,Er/NaYF4 upconversion nanoparticles (UCNPs) to generate a novel multifunctional nanoprobe, PEGylated UCNP@TEMPO@SiO2 for upconversion luminescence (UCL) and magnetic resonance dual-modality imaging. The electron spin resonance (ESR) signals generated by the TEMPO@SiO2 show an enhanced reduction resistance property for a period of time of up to 1 h, even in the presence of 5 mM ascorbic acid. The longitudinal relaxivity of PEGylated UCNPs@TEMPO@SiO2 nanocomposites is about 10 times stronger than that for free TEMPO radicals. The core-shell NaYF4:Yb,Er/NaYF4 UCNPs synthesized by this modified user-friendly one-pot solvothermal strategy show a significant enhancement of UCL emission of up to 60 times more than the core NaYF4:Yb,Er. Furthermore, the PEGylated UCNP@TEMPO@SiO2 nanocomposites were further used as multifunctional nanoprobes to explore their performance in the UCL imaging of living cells and T1-weighted MRI in vitro and in vivo.Nitroxide radicals, such as 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO) and its derivatives, have recently been used as contrast agents for magnetic resonance imaging (MRI) and electron

  1. Photoinhibition of photosynthesis in vivo results in singlet oxygen production detection via nitroxide-induced fluorescence quenching in broad bean leaves.

    PubMed

    Hideg, E; Kálai, T; Hideg, K; Vass, I

    1998-08-18

    In plants experiencing environmental stress, the formation of reactive oxygen is often presumed. In this study, singlet oxygen was detected in broad bean (Vicia faba) leaves that were photoinhibited in vivo. Detection was based on the reaction of singlet oxygen with DanePy (dansyl-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrole) yielding a nitroxide radical (DanePyO) which is EPR active and also features lower fluorescence compared to DanePy. The two (fluorescent and spin) sensor fuctions of DanePy are commensurate, which makes detecting singlet oxygen possible with a spectrofluorimeter in samples hard to measure with EPR spectroscopy [Kálai, T., Hideg, E., Vass, I., and Hideg, K. (1998) Free Radical Biol. Med. 24, 649-652]. We found that in leaves saturated with DanePy, the fluorescence of this double sensor was decreased when the leaves were photoinhibited by 1500 micromol m-2 s-1 photosynthetically active radiation. This fluorescence quenching is the first direct experimental evidence that photoinhibition of photosynthesis in vivo is accompanied by 1O2 production and is, at least partly, governed by the process characterized as acceptor side-induced photoinhibition in vitro.

  2. Increased oxidative modification of albumin when illuminated in vitro in the presence of a common sunscreen ingredient: protection by nitroxide radicals.

    PubMed

    Damiani, E; Carloni, P; Biondi, C; Greci, L

    2000-01-15

    We previously reported on the ability of dibenzoylmethane (DBM) and a relative, Parsol 1789, used as a ultraviolet A (UVA)-absorbing sunscreen, to generate free radicals upon illumination, and as a consequence, to inflict strand breaks in plasmid DNA in vitro. This study has now been extended to determine the effects of Parsol 1789 and DBM on proteins, under UVA illumination, with the sole purpose of gaining more knowledge on the photobiological effects of sunscreen chemicals. Parsol 1789 (100 microM) caused a 2-fold increase in protein carbonyl formation (an index of oxidative damage) in bovine serum albumin (BSA) when exposed to illumination, and this damage was both concentration- and time-dependent. The degree of protein damage was markedly reduced by the presence of free radical scavengers, namely piperidinic and indolinonic nitroxide radicals, in accordance with our previous study. Vitamin E had no effect under the conditions used. The results obtained corroborate the fact that Parsol 1789 generates free radicals upon illumination and that these are, most probably, responsible for the protein damage observed under the conditions used in our system. However, at present, we cannot extrapolate from these results the relevance to human use of sunscreens; therefore, further studies should be necessary to determine the efficacy at the molecular and cellular level of this UVA-absorber in order to ascertain protection against photocarcinogenic risk.

  3. Differential effects of the mitochondrial uncoupling agent, 2,4-dinitrophenol, or the nitroxide antioxidant, Tempol, on synaptic or nonsynaptic mitochondria after spinal cord injury.

    PubMed

    Patel, Samir P; Sullivan, Patrick G; Pandya, Jignesh D; Rabchevsky, Alexander G

    2009-01-01

    We recently documented the progressive nature of mitochondrial dysfunction over 24 hr after contusion spinal cord injury (SCI), but the underlying mechanism has not been elucidated. We investigated the effects of targeting two distinct possible mechanisms of mitochondrial dysfunction by using the mitochondrial uncoupler 2,4-dinitrophenol (2,4-DNP) or the nitroxide antioxidant Tempol after contusion SCI in rats. A novel aspect of this study was that all assessments were made in both synaptosomal (neuronal)- and nonsynaptosomal (glial and neuronal soma)-derived mitochondria 24 hr after injury. Mitochondrial uncouplers target Ca(2+) cycling and subsequent reactive oxygen species production in mitochondria after injury. When 2,4-DNP was injected 15 and 30 min after injury, mitochondrial function was preserved in both populations compared with vehicle-treated rats, whereas 1 hr postinjury treatment was ineffective. Conversely, targeting peroxynitrite with Tempol failed to maintain normal bioenergetics in synaptic mitochondria, but was effective in nonsynaptic mitochondria when administered 15 min after injury. When administered at 15 and 30 min after injury, increased hydroxynonenal, 3-NT, and protein carbonyl levels were significantly reduced by 2,4-DNP, whereas Tempol only reduced 3-NT and protein carbonyls after SCI. Despite such antioxidant effects, only 2,4-DNP was effective in preventing mitochondrial dysfunction, indicating that mitochondrial Ca(2+) overload may be the key mechanism involved in acute mitochondrial damage after SCI. Collectively, our observations demonstrate the significant role that mitochondrial dysfunction plays in SCI neuropathology. Moreover, they indicate that combinatorial therapeutic approaches targeting different populations of mitochondria holds great potential in fostering neuroprotection after acute SCI.

  4. Intraoral Mitochondrial-Targeted GS-Nitroxide, JP4-039, Radioprotects Normal Tissue in Tumor-Bearing Radiosensitive Fancd2(-/-) (C57BL/6) Mice.

    PubMed

    Shinde, Ashwin; Berhane, Hebist; Rhieu, Byung Han; Kalash, Ronny; Xu, Karen; Goff, Julie; Epperly, Michael W; Franicola, Darcy; Zhang, Xichen; Dixon, Tracy; Shields, Donna; Wang, Hong; Wipf, Peter; Parmar, Kalindi; Guinan, Eva; Kagan, Valerian; Tyurin, Vladimir; Ferris, Robert L; Zhang, Xiaolan; Li, Song; Greenberger, Joel S

    2016-02-01

    We evaluated normal tissue specific radioprotection of the oral cavity in radiosensitive Fanconi Anemia (FA) Fancd2(-/-) mice with orally established tumors using mitochondrial-targeted GS-nitroxide (JP4-039). Adult (10-12 weeks old) Fancd2(+/+), Fancd2(+/-) and Fancd2(-/-) mice (C57BL/6 background) and subgroups with orally established TC-1 epithelial cell tumors received a single fraction of 28 Gy or four daily fractions of 8 Gy to the head and neck. Subgroups received JP4-039 in F15 emulsion (F15/JP4-039; 0.4 mg/mouse), 4-amino-Tempo in F15 emulsion (F15/4-amino-Tempo; 0.2 mg/mouse) or F15 emulsion alone prior to each irradiation. Oral mucosa of Fancd2(-/-) mice showed baseline elevated RNA transcripts for Sod2, p53, p21 and Rad51 (all P < 0.0012) and suppressed levels of Nfkb and Tgfb, (all P < 0.0020) compared with Fancd2(+/+) mice. The oral mucosa in tumor-bearing mice of all genotypes showed decreased levels of p53 and elevated Tgfb and Gadd45a (P ≤ 0.0001 for all three genotypes). Intraoral F15/JP4-039, but not F15/4-amino-Tempo, modulated radiation-induced normal tissue transcript elevation, ameliorated mucosal ulceration and reduced the depletion of antioxidant stores in oral cavity tissue of all genotypes, but did not radioprotect tumors. Mitochondrial targeting makes F15/JP4-039 an effective normal tissue radioprotector for Fancd2(-/-) mice, as well as wild-type mice.

  5. Amyloid beta peptides do not form peptide-derived free radicals spontaneously, but can enhance metal-catalyzed oxidation of hydroxylamines to nitroxides.

    PubMed

    Dikalov, S I; Vitek, M P; Maples, K R; Mason, R P

    1999-04-02

    Amyloid beta (Abeta) peptides play an important role in the pathogenesis of Alzheimer's disease. Free radical generation by Abeta peptides was suggested to be a key mechanism of their neurotoxicity. Reports that neurotoxic free radicals derived from Abeta-(1-40) and Abeta-(25-35) peptides react with the spin trap N-tert-butyl-alpha-phenylnitrone (PBN) to form a PBN/.Abeta peptide radical adduct with a specific triplet ESR signal assert that the peptide itself was the source of free radicals. We now report that three Abeta peptides, Abeta-(1-40), Abeta-(25-35), and Abeta-(40-1), do not yield radical adducts with PBN from the Oklahoma Medical Research Foundation (OMRF). In contrast to OMRF PBN, incubation of Sigma PBN in phosphate buffer without Abeta peptides produced a three-line ESR spectrum. It was shown that this nitroxide is di-tert-butylnitroxide and is formed in the Sigma PBN solution as a result of transition metal-catalyzed auto-oxidation of the respective hydroxylamine present as an impurity in the Sigma PBN. Under some conditions, incubation of PBN from Sigma with Abeta-(1-40) or Abeta-(25-35) can stimulate the formation of di-tert-butylnitroxide. It was shown that Abeta peptides enhanced oxidation of cyclic hydroxylamine 1-hydroxy-4-oxo-2,2,6, 6-tetramethylpiperidine (TEMPONE-H), which was strongly inhibited by the treatment of phosphate buffer with Chelex-100. It was shown that ferric and cupric ions are effective oxidants of TEMPONE-H. The data obtained allow us to conclude that under some conditions toxic Abeta peptides Abeta-(1-40) and Abeta-(25-35) enhance metal-catalyzed oxidation of hydroxylamine derivatives, but do not spontaneously form peptide-derived free radicals.

  6. Mutant SOD1 microglia-generated nitroxidative stress promotes toxicity to human fetal neural stem cell-derived motor neurons through direct damage and noxious interactions with astrocytes

    PubMed Central

    Thonhoff, Jason R; Gao, Junling; Dunn, Tiffany J; Ojeda, Luis; Wu, Ping

    2012-01-01

    Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease. Human neural stem cells (hNSCs) may have the potential to replace lost motor neurons. The therapeutic efficacy of stem cell therapy depends greatly on the survival of grafted stem cell-derived motor neurons in the microenvironment of the spinal cord in ALS. After transplantation of hNSCs into the spinal cords of transgenic ALS rats, morphological analysis reveals that grafted hNSCs differentiate into motor neurons. However, hNSCs degenerate and show signs of nitroxidative damage at the disease end-stage. Using an in vitro coculture system, we systematically assess interactions between microglia and astroglia derived from both nontransgenic rats and transgenic rats expressing human mutant SOD1G93A before and after symptomatic disease onset, and determine the effects of such microglia-astroglia interactions on the survival of hNSC-derived motor neurons. We found that ALS microglia, specifically isolated after symptomatic disease onset, are directly toxic to hNSC-derived motor neurons. Furthermore, nontransgenic astrocytes not only lose their protective role in hNSC-derived motor neuron survival in vitro, but also exhibit toxic features when cocultured with mutant SOD1G93A microglia. Using inhibitors of inducible nitric oxide synthase and NADPH oxidase, we show that microglia-generated nitric oxide and superoxide partially contribute to motor neuron loss and astrocyte dysfunction in this coculture paradigm. In summary, reactive oxygen/nitrogen species released from overactivated microglia in ALS directly eliminate human neural stem cell-derived motor neurons and reduce the neuroprotective capacities of astrocytes PMID:23671793

  7. Synergistic induction of apoptosis and caspase-independent autophagic cell death by a combination of nitroxide Tempo and heat shock in human leukemia U937 cells.

    PubMed

    Zhao, Qing-Li; Fujiwara, Yoshisada; Kondo, Takashi

    2010-10-01

    We have shown that heat stress or a superoxide dismutase mimic nitroxide, Tempo, induces apoptosis, while their combination causes nonapoptotic cell death; however, the underlying mechanism for this switch remains unclear. Here we identified for the first time that 10 mM Tempo present during heating at 44°C for 30 min rapidly induced autophagy in U937 leukemic cells in spite of Bax activation and mitochondrial outer membrane (MOM) permeabilization. This co-treatment inhibited the processing of heat-activated procaspases-2, -8, -9 and -3 into active small subunits, leading to the inhibition of caspase-dependent apoptosis, and instead caused the induction of autophagy. The inactivation of caspases, a key event, could result from oxidation of active-site-CysSH of all caspases by a prooxidant oxo-ammonium cation, an intermediate derived Tempo during dismutation of heat-induced superoxide anion. In addition, the co-treatment caused mitochondrial calcium overloads, the mitochondrial inner membrane permeabilization, profound mitochondrial dysfunction, and liberation of Beclin 1 from the Bcl-2/Beclin 1 complex, all of which contributed to induction of autophagy. These autophagic cells underwent propidium iodide-positive necrosis in a delayed fashion, leading to the complete proliferative inhibition. Remarkably, ruthenium red and BAPTA, which interfere with mitochondrial calcium uptake, facilitated autophagic necrotic death. Cyclosporin A, which binds to cyclophilin D, had a similar necrotic effect. 3-Methyladenine facilitated the necrosis of autophagic cells. In contrast, 5 mM Tempo-44°C/10 min or 44°C/30 min induced Bax-mediated MOM permeabilization and caspase-dependent apoptosis more potently than Tempo alone. Thus, Tempo is a unique thermosensitizer to synergistically induce apoptosis and autophagic cell death.

  8. Real-time monitoring of drug-induced changes in the stomach acidity of living rats using improved pH-sensitive nitroxides and low-field EPR techniques

    NASA Astrophysics Data System (ADS)

    Potapenko, Dmitrii I.; Foster, Margaret A.; Lurie, David J.; Kirilyuk, Igor A.; Hutchison, James M. S.; Grigor'ev, Igor A.; Bagryanskaya, Elena G.; Khramtsov, Valery V.

    2006-09-01

    New improved pH-sensitive nitroxides were applied for in vivo studies. An increased stability of the probes towards reduction was achieved by the introduction of the bulky ethyl groups in the vicinity of the paramagnetic N sbnd O fragment. In addition, the range of pH sensitivity of the approach was extended by the synthesis of probes with two ionizable groups, and, therefore, with two p Ka values. Stability towards reduction and spectral characteristics of the three new probes were determined in vitro using 290 MHz radiofrequency (RF)- and X-band electron paramagnetic resonance (EPR), longitudinally detected EPR (LODEPR), and field-cycled dynamic nuclear polarization (FC-DNP) techniques. The newly synthesized probe, 4-[bis(2-hydroxyethyl)amino]-2-pyridine-4-yl-2,5,5-triethyl-2,5-dihydro-1 H-imidazol-oxyl, was found to be the most appropriate for the application in the stomach due to both higher stability and convenient pH sensitivity range from pH 1.8 to 6. LODEPR, FC-DNP and proton-electron double resonance imaging (PEDRI) techniques were used to detect the nitroxide localization and acidity in the rat stomach. Improved probe characteristics allowed us to follow in vivo the drug-induced perturbation in the stomach acidity and its normalization afterwards during 1 h or longer period of time. The results show the applicability of the techniques for monitoring drug pharmacology and disease in the living animals.

  9. Dynamics of 4-oxo-TEMPO-d16-15N nitroxide-propylene glycol system studied by ESR and ESE in liquid and glassy state in temperature range 10-295 K

    NASA Astrophysics Data System (ADS)

    Goslar, Janina; Hoffmann, Stanislaw K.; Lijewski, Stefan

    2016-08-01

    ESR spectra and electron spin relaxation of nitroxide radical in 4-oxo-TEMPO-d16-15N in propylene glycol were studied at X-band in the temperature range 10-295 K. The spin-lattice relaxation in the liquid viscous state determined from the resonance line shape is governed by three mechanisms occurring during isotropic molecular reorientations. In the glassy state below 200 K the spin-lattice relaxation, phase relaxation and electron spin echo envelope modulations (ESEEM) were studied by pulse spin echo technique using 2-pulse and 3-pulse induced signals. Electron spin-lattice relaxation is governed by a single non-phonon relaxation process produced by localized oscillators of energy 76 cm-1. Electron spin dephasing is dominated by a molecular motion producing a resonance-type peak in the temperature dependence of the dephasing rate around 120 K. The origin of the peak is discussed and a simple method for the peak shape analysis is proposed, which gives the activation energy of a thermally activated motion Ea = 7.8 kJ/mol and correlation time τ0 = 10-8 s. The spin echo amplitude is strongly modulated and FT spectrum contains a doublet of lines centered around the 2D nuclei Zeeman frequency. The splitting into the doublet is discussed as due to a weak hyperfine coupling of nitroxide unpaired electron with deuterium of reorienting CD3 groups.

  10. Heisenberg spin exchange effects of nitroxide radicals on Overhauser dynamic nuclear polarization in the low field limit at 1.5mT.

    PubMed

    Lingwood, Mark D; Ivanov, Ivan A; Cote, Alissa R; Han, Songi

    2010-05-01

    Nuclear magnetic resonance (NMR) and magnetic resonance imaging (MRI) at very low magnetic fields (0.05-20mT) have gained interest due to the simple and portable magnet design and newly emerging applications outside of the usual laboratory setting. A method to enhance the NMR signal is needed due to the low thermal polarization of nuclear spins at these fields; dynamic nuclear polarization (DNP) via the Overhauser effect from free radicals is an attractive option. In this report we describe a DNP-enhanced NMR system operating at a fixed field of 1.5mT and measure (1)H signal enhancements of up to -350 fold during the saturation of a selected electron spin resonance (ESR) transition of dissolved nitroxide radicals. This -350 fold enhanced polarization is equivalent to what would be obtained by prepolarization in a 0.53T field. The ESR spectra at varying radical concentrations are indirectly found through DNP-enhanced NMR detection. Here, ESR line broadening at higher radical concentrations due to Heisenberg electron spin exchange is observed. Enhancements in the limit of maximum power are reported as a function of concentration for three ESR transitions, and are found to increase with concentration. The >300 fold (1)H NMR signal amplifications achievable at 1.5mT will reduce experimental time by several orders of magnitude, permitting NMR relaxation, imaging or pulsed-field gradient diffusion experiments that are inaccessible without using the DNP effect at 1.5mT. We demonstrate the potential benefit of such large signal amplification schemes through T(1) and T(2) relaxation measurements carried out in a much shorter time when employing DNP. Finally, we compare our results to those obtained in the earth's magnetic field and find that the signal to noise ratio (SNR) of DNP-enhanced signal at 1.5mT is much greater than that obtained by previous studies utilizing DNP enhancement in the 0.05mT earth's magnetic field. Copyright 2010 Elsevier Inc. All rights reserved.

  11. Effect of gallium-porphyrin analogue ATX-70 on nitroxide formation from a cyclic secondary amine by ultrasound: On the mechanism of sonodynamic activation

    SciTech Connect

    Miyoshi, N.; Misik, V.; Riesz, P.

    1995-08-01

    Sonodynamic therapy is a promising new modality for cancer treatment based on the synergistic effect on tumor cell killing by combination of a drug (typically a photosensitizer) and ultra-sound. The mechanism of sonodynamic action was suggested to involve photoexcitation of the sensitizer by sonoluminescent light, with subsequent formation of singlet oxygen. In this work we studied the aqueous sonsochemical reactions of the gallium-porphyrin derivative ATX-70, one of the most active sonodynamic agents found, using 50 kHz ultrasound. The experiments were carried out in the presence of 2,2,6,6-tetramethyl-4-piperidone hydrochloride (TMP), which reacts with singlet oxygen or {degrees}OH radicals to give the EPR-detectable nitroxide 2,2,6,6-tetramethyl-piperidone-N-oxyl (TMP-NO). Recently it has been suggested that the enhancement of TMP-NO yields in the presence of aqueous solutions of ATX-70 exposed to ultrasound was evidence for the formation of singlet oxygen in the system. Our results show that the surfactant cetyltrimethylammonium bromide (CTAB) can mimic the ATX-70-induced increase in the TMP-NO signal, but it fails to reproduce the behavior of ATX-70 in D{sub 2}O: while the yields of TMP-NO in the presence of ATX-70 increase in D{sub 2}O, the opposite effect was found with the surfactant CTAB. However, our data show that the increased TMP-NO yields in D{sub 2}O are paralleled by an increased concentration of ATX-70 dimer, a form that is inactive in the photochemical generation of singlet oxygen. Finding the ATX-70-dependent enhancement of the TMP-NO signal was highest at {approximately} 20% O{sub 2}, in both N{sub 2}/O{sub 2} and argon/O{sub 2} mixtures, and decreased with increasing oxygen concentration is not compatible with the singlet oxygen mechanism. Finally, results on the temperature dependence of the ATX-70-induced formation of TMP-NO are not consistent with the photochemical excitation of ATX-70 by sonoluminescent light. 44 refs., 10 figs., 1 tab.

  12. DFT study of nitroxide radicals: explicit modeling of solvent effects on the structural and electronic characteristics of 4-amino-2,2,6,6-tetramethyl-piperidine-N-oxyl.

    PubMed

    Ikryannikova, Larissa N; Ustynyuk, Leila Yu; Tikhonov, Alexander N

    2010-05-01

    An explicit DFT modeling of water surroundings on the electron paramagnetic resonance properties of 4-amino-2,2,6,6-tetramethyl-piperidine-N-oxyl (TA) has been performed. A stepwise hydration of TA is accompanied with certain changes in geometrical parameters (bond lengths and angles) and redistribution of partial electric charges in TA. An aqueous cluster of 45 water molecules can be considered as an appropriate model for a complete aqueous shell around TA, although most of the structural and electronic characteristics of TA already converge at about 10 water molecules. Water surroundings induce an increase in electron spin density on the nitrogen atom of the nitroxide fragment due to stabilization of the polar resonance structure > N(+*)-O(-) at the expense of less polar structure > N-O*. The water-induced rise of the isotropic splitting constant a(iso), calculated from the contact term of the hyperfine interaction, comprises Deltaa(iso)(rho(N2)) = 2.2-2.5 G, which is typical of experimental value for TA. There are two contributions to the solvent effect on the a(iso)(rho(N2)) value: the redistribution of spin density in the nitroxide fragment (polarity effect) and water-induced distortions of TA geometry. Microscopic variations in a hydrogen-bonded water network cause noticeable fluctuations of the splitting constant a(iso)(rho(N2)). Calculations of the atomic spin density (sigma(N2)) allowed us to compute the splitting constant from the relationship a(iso)(sigma(N2)) = Qsigma(N2), where Q = 36.2 G. A practical advantage of using this relationship is that it gives 'smoothed' values of the splitting constant, which are sensitive to the environment polarity but remain tolerant to microscopic fluctuations of the hydrogen-bonded water network around a spin-label molecule.

  13. Dynamic changes in the distribution and time course of blood-brain barrier-permeative nitroxides in the mouse head with EPR imaging: visualization of blood flow in a mouse model of ischemia.

    PubMed

    Emoto, Miho C; Sato-Akaba, Hideo; Hirata, Hiroshi; Fujii, Hirotada G

    2014-09-01

    Electron paramagnetic resonance (EPR) imaging using nitroxides as redox-sensitive probes is a powerful, noninvasive method that can be used under various physiological conditions to visualize changes in redox status that result from oxidative damage. Two blood-brain barrier-permeative nitroxides, 3-hydroxymethyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (HMP) and 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-yloxy (MCP), have been widely used as redox-sensitive probes in the brains of small animals, but their in vivo distribution and properties have not yet been analyzed in detail. In this study, a custom-made continuous-wave three-dimensional (3D) EPR imager was used to obtain 3D EPR images of mouse heads using MCP or HMP. This EPR imager made it possible to take 3D EPR images reconstructed from data from 181 projections acquired every 60s. Using this improved EPR imager and magnetic resonance imaging, the distribution and reduction time courses of HMP and MCP were examined in mouse heads. EPR images of living mice revealed that HMP and MCP have different distributions and different time courses for entering the brain. Based on the pharmacokinetics of the reduction reactions of HMP and MCP in the mouse head, the half-lives of HMP and MCP were clearly and accurately mapped pixel by pixel. An ischemic mouse model was prepared, and the half-life of MCP was mapped in the mouse head. Compared to the half-life in control mice, the half-life of MCP in the ischemic model mouse brain was significantly increased, suggesting a shift in the redox balance. This in vivo EPR imaging method using BBB-permeative MCP is a useful noninvasive method for assessing changes in the redox status in mouse brains under oxidative stress. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Heterologous expression of mitochondria-targeted microbial nitrilase enzymes increases cyanide tolerance in Arabidopsis.

    PubMed

    Molojwane, E; Adams, N; Sweetlove, L J; Ingle, R A

    2015-07-01

    Anthropogenic activities have resulted in cyanide (CN) contamination of both soil and water in many areas of the globe. While plants possess a detoxification pathway that serves to degrade endogenously generated CN, this system is readily overwhelmed, limiting the use of plants in bioremediation. Genetic engineering of additional CN degradation pathways in plants is one potential strategy to increase their tolerance to CN. Here we show that heterologous expression of microbial nitrilase enzymes targeted to the mitochondria increases CN tolerance in Arabidopsis. Root length in seedlings expressing either a CN dihydratase from Bacillus pumilis or a CN hydratase from Neurospora crassa was increased by 45% relative in wild-type plants in the presence of 50 μm KCN. We also demonstrate that in contrast to its strong inhibitory effects on seedling establishment, seed germination of the Col-0 ecotype of Arabidopsis is unaffected by CN. © 2015 German Botanical Society and The Royal Botanical Society of the Netherlands.

  15. A graphene oxide based smart drug delivery system for tumor mitochondria-targeting photodynamic therapy

    NASA Astrophysics Data System (ADS)

    Wei, Yanchun; Zhou, Feifan; Zhang, Da; Chen, Qun; Xing, Da

    2016-02-01

    Subcellular organelles play critical roles in cell survival. In this work, a novel photodynamic therapy (PDT) drug delivery and phototoxicity on/off nano-system based on graphene oxide (NGO) as the carrier is developed to implement subcellular targeting and attacking. To construct the nanodrug (PPa-NGO-mAb), NGO is modified with the integrin αvβ3 monoclonal antibody (mAb) for tumor targeting. Pyropheophorbide-a (PPa) conjugated with polyethylene-glycol is used to cover the surface of the NGO to induce phototoxicity. Polyethylene-glycol phospholipid is loaded to enhance water solubility. The results show that the phototoxicity of PPa on NGO can be switched on and off in organic and aqueous environments, respectively. The PPa-NGO-mAb assembly is able to effectively target the αvβ3-positive tumor cells with surface ligand and receptor recognition; once endocytosized by the cells, they are observed escaping from lysosomes and subsequently transferring to the mitochondria. In the mitochondria, the `on' state PPa-NGO-mAb performs its effective phototoxicity to kill cells. The biological and physical dual selections and on/off control of PPa-NGO-mAb significantly enhance mitochondria-mediated apoptosis of PDT. This smart system offers a potential alternative to drug delivery systems for cancer therapy.Subcellular organelles play critical roles in cell survival. In this work, a novel photodynamic therapy (PDT) drug delivery and phototoxicity on/off nano-system based on graphene oxide (NGO) as the carrier is developed to implement subcellular targeting and attacking. To construct the nanodrug (PPa-NGO-mAb), NGO is modified with the integrin αvβ3 monoclonal antibody (mAb) for tumor targeting. Pyropheophorbide-a (PPa) conjugated with polyethylene-glycol is used to cover the surface of the NGO to induce phototoxicity. Polyethylene-glycol phospholipid is loaded to enhance water solubility. The results show that the phototoxicity of PPa on NGO can be switched on and off in organic and aqueous environments, respectively. The PPa-NGO-mAb assembly is able to effectively target the αvβ3-positive tumor cells with surface ligand and receptor recognition; once endocytosized by the cells, they are observed escaping from lysosomes and subsequently transferring to the mitochondria. In the mitochondria, the `on' state PPa-NGO-mAb performs its effective phototoxicity to kill cells. The biological and physical dual selections and on/off control of PPa-NGO-mAb significantly enhance mitochondria-mediated apoptosis of PDT. This smart system offers a potential alternative to drug delivery systems for cancer therapy. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr07785k

  16. Mitochondrial Dysfunction in the Pathogenesis of Rett Syndrome: Implications for Mitochondria-Targeted Therapies

    PubMed Central

    Shulyakova, Natalya; Andreazza, Ana C.; Mills, Linda R.; Eubanks, James H.

    2017-01-01

    First described over 50 years ago, Rett syndrome (RTT) is a neurodevelopmental disorder caused primarily by mutations of the X-linked MECP2 gene. RTT affects predominantly females, and has a prevalence of roughly 1 in every 10,000 female births. Prior to the discovery that mutations of MECP2 are the leading cause of RTT, there were suggestions that RTT could be a mitochondrial disease. In fact, several reports documented altered mitochondrial structure, and deficiencies in mitochondrial enzyme activity in different cells or tissues derived from RTT patients. With the identification of MECP2 as the causal gene, interest largely shifted toward defining the normal function of MeCP2 in the brain, and how its absence affects the neurodevelopment and neurophysiology. Recently, though, interest in studying mitochondrial function in RTT has been reignited, at least in part due to observations suggesting systemic oxidative stress does play a contributing role in RTT pathogenesis. Here we review data relating to mitochondrial alterations at the structural and functional levels in RTT patients and model systems, and present a hypothesis for how the absence of MeCP2 could lead to altered mitochondrial function and elevated levels of cellular oxidative stress. Finally, we discuss the prospects for treating RTT using interventions that target specific aspects of mitochondrial dysfunction and/or oxidative stress. PMID:28352216

  17. Oxidative stress and mitochondrial dysfunction in atherosclerosis: mitochondria-targeted antioxidants as potential therapy.

    PubMed

    Victor, V M; Apostolova, N; Herance, R; Hernandez-Mijares, A; Rocha, M

    2009-01-01

    Chronic and acute overproduction of reactive oxygen species (ROS) under pathophysiologic conditions forms an integral part of the development of cardiovascular diseases (CVD), and in particular atherosclerosis. These ROS are released from different sources, such as xanthine oxidase, lipoxygenase, nicotinamide adenine dinucleotide phosphate oxidase, the uncoupling of nitric oxide synthase and, in particular, mitochondria. Endothelial dysfunction, characterized by a loss of nitric oxide (NO) bioactivity, occurs early on in the development of atherosclerosis, and determines future vascular complications. Although the molecular mechanisms responsible for mitochondria-mediated disease processes are not clear, oxidative stress seems to play an important role. In general, ROS are essential to cell function, but adequate levels of antioxidant defenses are required in order to avoid the harmful effects of excessive ROS production. Mitochondrial oxidative stress damage and dysfunction contribute to a number of cell pathologies that manifest themselves through a range of conditions. This review considers the process of atherosclerosis from a mitochondrial perspective, and assesses strategies for the targeted delivery of antioxidants to mitochondria that are currently under development. We will provide a summary of the following areas: the cellular metabolism of reactive oxygen species (ROS) and its role in pathophysiological processes such as atherosclerosis; currently available antioxidants and possible reasons for their efficacy and inefficacy in ameliorating oxidative stress-mediated diseases; and recent developments in mitochondrially-targeted antioxidants that concentrate on the matrix-facing surface of the inner mitochondrial membrane in order to protect against mitochondrial oxidative damage, and their therapeutic potential as a treatment for atherosclerosis.

  18. Ester-Modified Cyclometalated Iridium(III) Complexes as Mitochondria-Targeting Anticancer Agents

    PubMed Central

    Wang, Fang-Xin; Chen, Mu-He; Hu, Xiao-Ying; Ye, Rui-Rong; Tan, Cai-Ping; Ji, Liang-Nian; Mao, Zong-Wan

    2016-01-01

    Organometallic iridium complexes are potent anticancer candidates which act through different mechanisms from cisplatin-based chemotherapy regimens. Here, ten phosphorescent cyclometalated iridium(III) complexes containing 2,2′-bipyridine-4,4′-dicarboxylic acid and its diester derivatives as ligands are designed and synthesized. The modification by ester group, which can be hydrolysed by esterase, facilitates the adjustment of drug-like properties. The quantum yields and emission lifetimes are influenced by variation of the ester substituents on the Ir(III) complexes. The cytotoxicity of these Ir(III) complexes is correlated with the length of their ester groups. Among them, 4a and 4b are found to be highly active against a panel of cancer cells screened, including cisplatin-resistant cancer cells. Mechanism studies in vitro indicate that they undergo hydrolysis of ester bonds, accumulate in mitochondria, and induce a series of cell-death related events mediated by mitochondria. Furthermore, 4a and 4b can induce pro-death autophagy and apoptosis simultaneously. Our study indicates that ester modification is a simple and feasible strategy to enhance the anticancer potency of Ir(III) complexes. PMID:27958338

  19. Evaluation of respiration of mitochondria in cancer cells exposed to mitochondria-targeted agents.

    PubMed

    Kluckova, Katarina; Dong, Lan-Feng; Bajzikova, Martina; Rohlena, Jakub; Neuzil, Jiri

    2015-01-01

    Respiration is one of the major functions of mitochondria, whereby these vital organelles use oxygen to produce energy. Many agents that may be of potential clinical relevance act by targeting mitochondria, where they may suppress mitochondrial respiration. It is therefore important to evaluate this process and understand how this is modulated by small molecules. Here, we describe the general methodology to assess respiration in cultured cells, followed by the evaluation of the effect of one anticancer agent targeted to mitochondria on this process, and also how to assess this in tumor tissue.

  20. Melatonin as a mitochondria-targeted antioxidant: one of evolution's best ideas.

    PubMed

    Reiter, Russel J; Rosales-Corral, Sergio; Tan, Dun Xian; Jou, Mei Jie; Galano, Annia; Xu, Bing

    2017-09-01

    Melatonin is an ancient antioxidant. After its initial development in bacteria, it has been retained throughout evolution such that it may be or may have been present in every species that have existed. Even though it has been maintained throughout evolution during the diversification of species, melatonin's chemical structure has never changed; thus, the melatonin present in currently living humans is identical to that present in cyanobacteria that have existed on Earth for billions of years. Melatonin in the systemic circulation of mammals quickly disappears from the blood presumably due to its uptake by cells, particularly when they are under high oxidative stress conditions. The measurement of the subcellular distribution of melatonin has shown that the concentration of this indole in the mitochondria greatly exceeds that in the blood. Melatonin presumably enters mitochondria through oligopeptide transporters, PEPT1, and PEPT2. Thus, melatonin is specifically targeted to the mitochondria where it seems to function as an apex antioxidant. In addition to being taken up from the circulation, melatonin may be produced in the mitochondria as well. During evolution, mitochondria likely originated when melatonin-forming bacteria were engulfed as food by ancestral prokaryotes. Over time, engulfed bacteria evolved into mitochondria; this is known as the endosymbiotic theory of the origin of mitochondria. When they did so, the mitochondria retained the ability to synthesize melatonin. Thus, melatonin is not only taken up by mitochondria but these organelles, in addition to many other functions, also probably produce melatonin as well. Melatonin's high concentrations and multiple actions as an antioxidant provide potent antioxidant protection to these organelles which are exposed to abundant free radicals.

  1. Curcumin "Drug" Stabilized in Oxidovanadium(IV)-BODIPY Conjugates for Mitochondria-Targeted Photocytotoxicity.

    PubMed

    Bhattacharyya, Utso; Kumar, Brijesh; Garai, Aditya; Bhattacharyya, Arnab; Kumar, Arun; Banerjee, Samya; Kondaiah, Paturu; Chakravarty, Akhil R

    2017-10-03

    Ternary oxidovanadium(IV) complexes of curcumin (Hcur), dipicolylamine (dpa) base, and its derivatives having pendant noniodinated and di-iodinated boron-dipyrromethene (BODIPY) moiety (L1 and L2, respectively), namely, [VO(dpa)(cur)]ClO4 (1), [VO(L1)(cur)]ClO4 (2), and [VO(L2)(cur)]ClO4 (3) and their chloride salts (1a-3a) were prepared, characterized, and studied for anticancer activity. The chloride salts were used for biological studies due to their aqueous solubility. Complex 1 was structurally characterized by single-crystal X-ray crystallography. The complex has a VO(2+) moiety bound to dpa ligand showing N,N,N-coordination in a facial mode, and curcumin is bound in its mono-anionic enolic form. The V-O(cur) distances are 1.950(18) and 1.977(16) Å, while the V-N bond lengths are 2.090(2), 2.130(2), and 2.290(2) Å. The bond trans to V═O is long due to trans effect. The complexes are stable in a solution phase over a long period of time of 48 h without showing any apparent degradation of the curcumin ligand. The diiodo-BODIPY ligand (L2) or Hcur alone showed limited solution stability in dark. The emissive BODIPY (L1) containing complex 2a showed preferential mitochondrial localization in MCF-7 cells in cellular imaging experiments. The cytotoxicity of the complexes was studied by MTT assay. The BODIPY complex 3a showed excellent photodynamic therapy effect in visible light (400-700 nm) giving IC50 values of 2-6 μM in HeLa and MCF-7 cancer cells, while being less toxic in dark (∼100 μM). The cell death was apoptotic in nature involving reactive oxygen species (ROS). Mechanistic data from pUC19 DNA photocleavage studies revealed photogenerated ROS as primarily (1)O2 from the BODIPY moiety and ·OH radicals from the curcumin ligand.

  2. Purification and In Vitro Activity of Mitochondria Targeted Nitrogenase Cofactor Maturase NifB.

    PubMed

    Burén, Stefan; Jiang, Xi; López-Torrejón, Gema; Echavarri-Erasun, Carlos; Rubio, Luis M

    2017-01-01

    Active NifB is a milestone in the process of engineering nitrogen fixing plants. NifB is an extremely O2-sensitive S-adenosyl methionine (SAM)-radical enzyme that provides the key metal cluster intermediate (NifB-co) for the biosyntheses of the active-site cofactors of all three types of nitrogenases. NifB and NifB-co are unique to diazotrophic organisms. In this work, we have expressed synthetic codon-optimized versions of NifB from the γ-proteobacterium Azotobacter vinelandii and the thermophilic methanogen Methanocaldococcus infernus in Saccharomyces cerevisiae and in Nicotiana benthamiana. NifB proteins were targeted to the mitochondria, where O2 consumption is high and bacterial-like [Fe-S] cluster assembly operates. In yeast, NifB proteins were co-expressed with NifU, NifS, and FdxN proteins that are involved in NifB [Fe-S] cluster assembly and activity. The synthetic version of thermophilic NifB accumulated in soluble form within the yeast cell, while the A. vinelandii version appeared to form aggregates. Similarly, NifB from M. infernus was expressed at higher levels in leaves of Nicotiana benthamiana and accumulated as a soluble protein while A. vinelandii NifB was mainly associated with the non-soluble cell fraction. Soluble M. infernus NifB was purified from aerobically grown yeast and biochemically characterized. The purified protein was functional in the in vitro FeMo-co synthesis assay. This work presents the first active NifB protein purified from a eukaryotic cell, and highlights the importance of screening nif genes from different organisms in order to sort the best candidates to assemble a functional plant nitrogenase.

  3. Monofunctional BODIPY-Appended Imidazoplatin for Cellular Imaging and Mitochondria-Targeted Photocytotoxicity.

    PubMed

    Raza, Md Kausar; Gautam, Srishti; Garai, Aditya; Mitra, Koushambi; Kondaiah, Paturu; Chakravarty, Akhil R

    2017-09-18

    Monofunctional platinum(II) complexes of formulation cis-[Pt(NH3)2(L)Cl](NO3), where L is an imidazole base conjugated to 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) with emissive (L(1) in 1) and nonemissive (L(2) in 2) moieties were prepared and characterized, and their singlet oxygen-mediated photoinduced cytotoxicity was studied. The 1-methylimidazole (1-MeIm) complex 3 was prepared as a control and for structural characterization by X-ray crystallography. Complexes 1 and 2 showed strong visible absorption bands at 500 nm (ε = 2.7 × 10(4) M(-1) cm(-1)) and 540 nm (1.4 × 10(4) M(-1) cm(-1)). Complex 1 is emissive with a band at 510 nm (ΦF = 0.09) in 1% dimethyl sulfoxide/Dulbecco's Modified Eagle's Medium (pH 7.2). Singlet oxygen generation upon photoirradiation with visible light (400-700 nm) was evidenced from 1,3-diphenylisobenzofuran titration experiments showing significant photosensitizing ability of the BODIPY complexes. Both 1 and 2 were remarkably photocytotoxic in visible light (400-700 nm, 10 J cm(-2)) in skin keratinocyte HaCaT and breast cancer MCF-7 cells giving IC50 values in nanomolar concentration. The complexes were, however, essentially nontoxic to the cells in the dark (IC50 > 80 μM). Complex 2 having a diiodo-BODIPY unit is nonemissive but an efficient photosensitizer with high singlet oxygen generation ability in visible light (400-700 nm). Confocal microscopy using the emissive complex 1 showed significant mitochondrial localization of the complex. Cell death via apoptotic pathway was observed from the Annexin-V-FITC/PI assay. The formation of Pt-DNA adducts was evidenced from the binding experiments of the complexes 1 and 2 with 9-ethylguanine as a model nucleobase from (1)H NMR and mass spectral studies.

  4. Mitochondria-targeted dodecyltriphenylphosphonium (C12TPP) combats high-fat-diet-induced obesity in mice

    PubMed Central

    Kalinovich, A V; Mattsson, C L; Youssef, M R; Petrovic, N; Ost, M; Skulachev, V P; Shabalina, I G

    2016-01-01

    Background: A membrane-penetrating cation, dodecyltriphenylphosphonium (C12TPP), facilitates the recycling of fatty acids in the artificial lipid membrane and mitochondria. C12TPP can dissipate mitochondrial membrane potential and may affect total energy expenditure and body weight in animals and humans. Methods: We investigated the metabolic effects of C12TPP in isolated brown-fat mitochondria, brown adipocyte cultures and mice in vivo. Experimental approaches included the measurement of oxygen consumption, carbon dioxide production, western blotting, magnetic resonance imaging and bomb calorimetry. Results: In mice, C12TPP (50 μmol per (day•kg body weight)) in the drinking water significantly reduced body weight (12%, P<0.001) and body fat mass (24%, P<0.001) during the first 7 days of treatment. C12TPP did not affect water palatability and intake or the energy and lipid content in feces. The addition of C12TPP to isolated brown-fat mitochondria resulted in increased oxygen consumption. Three hours of pretreatment with C12TPP also increased oligomycin-insensitive oxygen consumption in brown adipocyte cultures (P<0.01). The effects of C12TPP on mitochondria, cells and mice were independent of uncoupling protein 1 (UCP1). However, C12TPP treatment increased the mitochondrial protein levels in the brown adipose tissue of both wild-type and UCP1-knockout mice. Pair-feeding revealed that one-third of the body weight loss in C12TPP-treated mice was due to reduced food intake. C12TPP treatment elevated the resting metabolic rate (RMR) by up to 18% (P<0.05) compared with pair-fed animals. C12TPP reduced the respiratory exchange ratio, indicating enhanced fatty acid oxidation in mice. Conclusions: C12TPP combats diet-induced obesity by reducing food intake, increasing the RMR and enhancing fatty acid oxidation. PMID:27534841

  5. Oxidative Stress and Mitochondrial Dysfunction across Broad-Ranging Pathologies: Toward Mitochondria-Targeted Clinical Strategies

    PubMed Central

    d'Ischia, Marco; Gadaleta, Maria Nicola; Pallardó, Federico V.; Petrović, Sandra; Tiano, Luca; Zatterale, Adriana

    2014-01-01

    Beyond the disorders recognized as mitochondrial diseases, abnormalities in function and/or ultrastructure of mitochondria have been reported in several unrelated pathologies. These encompass ageing, malformations, and a number of genetic or acquired diseases, as diabetes and cardiologic, haematologic, organ-specific (e.g., eye or liver), neurologic and psychiatric, autoimmune, and dermatologic disorders. The mechanistic grounds for mitochondrial dysfunction (MDF) along with the occurrence of oxidative stress (OS) have been investigated within the pathogenesis of individual disorders or in groups of interrelated disorders. We attempt to review broad-ranging pathologies that involve mitochondrial-specific deficiencies or rely on cytosol-derived prooxidant states or on autoimmune-induced mitochondrial damage. The established knowledge in these subjects warrants studies aimed at elucidating several open questions that are highlighted in the present review. The relevance of OS and MDF in different pathologies may establish the grounds for chemoprevention trials aimed at compensating OS/MDF by means of antioxidants and mitochondrial nutrients. PMID:24876913

  6. Cancer Cell Mitochondria Targeting by Pancratistatin Analogs is Dependent on Functional Complex II and III

    PubMed Central

    Ma, Dennis; Pignanelli, Christopher; Tarade, Daniel; Gilbert, Tyler; Noel, Megan; Mansour, Fadi; Adams, Scott; Dowhayko, Alexander; Stokes, Kyle; Vshyvenko, Sergey; Hudlicky, Tomas; McNulty, James; Pandey, Siyaram

    2017-01-01

    Enhanced mitochondrial stability and decreased dependence on oxidative phosphorylation confer an acquired resistance to apoptosis in cancer cells, but may present opportunities for therapeutic intervention. The compound pancratistatin (PST) has been shown to selectively induce apoptosis in cancer cells. However, its low availability in nature has hindered its clinical advancement. We synthesized PST analogs and a medium-throughput screen was completed. Analogs SVTH-7, -6, and -5 demonstrated potent anti-cancer activity greater than PST and several standard chemotherapeutics. They disrupted mitochondrial function, activated the intrinsic apoptotic pathway, and reduced growth of tumor xenografts in vivo. Interestingly, the pro-apoptotic effects of SVTH-7 on cancer cells and mitochondria were abrogated with the inhibition of mitochondrial complex II and III, suggesting mitochondrial or metabolic vulnerabilities may be exploited by this analog. This work provides a scaffold for characterizing distinct mitochondrial and metabolic features of cancer cells and reveals several lead compounds with high therapeutic potential. PMID:28220885

  7. Cancer Cell Mitochondria Targeting by Pancratistatin Analogs is Dependent on Functional Complex II and III.

    PubMed

    Ma, Dennis; Pignanelli, Christopher; Tarade, Daniel; Gilbert, Tyler; Noel, Megan; Mansour, Fadi; Adams, Scott; Dowhayko, Alexander; Stokes, Kyle; Vshyvenko, Sergey; Hudlicky, Tomas; McNulty, James; Pandey, Siyaram

    2017-02-21

    Enhanced mitochondrial stability and decreased dependence on oxidative phosphorylation confer an acquired resistance to apoptosis in cancer cells, but may present opportunities for therapeutic intervention. The compound pancratistatin (PST) has been shown to selectively induce apoptosis in cancer cells. However, its low availability in nature has hindered its clinical advancement. We synthesized PST analogs and a medium-throughput screen was completed. Analogs SVTH-7, -6, and -5 demonstrated potent anti-cancer activity greater than PST and several standard chemotherapeutics. They disrupted mitochondrial function, activated the intrinsic apoptotic pathway, and reduced growth of tumor xenografts in vivo. Interestingly, the pro-apoptotic effects of SVTH-7 on cancer cells and mitochondria were abrogated with the inhibition of mitochondrial complex II and III, suggesting mitochondrial or metabolic vulnerabilities may be exploited by this analog. This work provides a scaffold for characterizing distinct mitochondrial and metabolic features of cancer cells and reveals several lead compounds with high therapeutic potential.

  8. Mitochondria-Targeted Antioxidant Mitoquinone Reduces Cisplatin-Induced Ototoxicity in Guinea Pigs.

    PubMed

    Tate, Alan D; Antonelli, Patrick J; Hannabass, Kyle R; Dirain, Carolyn O

    2017-03-01

    Objective To determine if mitoquinone (MitoQ) attenuates cisplatin-induced hearing loss in guinea pigs. Study Design Prospective and controlled animal study. Setting Academic, tertiary medical center. Subjects and Methods Guinea pigs were injected subcutaneously with either 5 mg/kg MitoQ (n = 9) or normal saline (control, n = 9) for 7 days and 1 hour before receiving a single dose of 10 mg/kg cisplatin. Auditory brainstem response thresholds were measured before MitoQ or saline administration and 3 to 4 days after cisplatin administration. Results Auditory brainstem response threshold shifts after cisplatin treatment were smaller by 28 to 47 dB in guinea pigs injected with MitoQ compared with those in the control group at all tested frequencies (4, 8, 16, and 24 kHz, P = .0002 to .04). Scanning electron microscopy of cochlear hair cells showed less outer hair cell loss and damage in the MitoQ group. Conclusion MitoQ reduced cisplatin-induced hearing loss in guinea pigs. MitoQ appears worthy of further investigation as a means of preventing cisplatin ototoxicity in humans.

  9. Cyclometalated iridium(III) complexes as mitochondria-targeted anticancer agents.

    PubMed

    Xiong, Kai; Chen, Yu; Ouyang, Cheng; Guan, Rui-Lin; Ji, Liang-Nian; Chao, Hui

    2016-06-01

    Four cyclometalated iridium(III) complexes [Ir(dfppy)2(L)](+) (dfppy = 2-(2,4-difluorophenyl)pyridine, L = 6-(pyridin-2-yl)-1,3,5-triazine-2,4-diamine, Ir1; 6-(isoquinolin-1-yl)-1,3,5-triazine-2,4-diamine, Ir2; 6-(quinolin-2-yl)-1,3,5-triazine-2,4-diamine, Ir3; 6-(isoquinolin-3-yl)-1,3,5-triazine-2,4-diamine, Ir4) have been synthesized and characterized. Distinct from cisplatin, Ir1-Ir4 could specifically target mitochondria and induced apoptosis against various cancer cell lines, especially for cisplatin resistant cells. ICP-MS results indicated that Ir1-Ir4 were taken up via different mechanism for cancer cells and normal cells, which resulted in their high selectivity. The structure-activity relationship and signaling pathways were also discussed. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  10. Cyclometalated iridium(III)-guanidinium complexes as mitochondria-targeted anticancer agents.

    PubMed

    Song, Xing-Dong; Kong, Xia; He, Shu-Fen; Chen, Jia-Xi; Sun, Jing; Chen, Bing-Bing; Zhao, Jin-Wu; Mao, Zong-Wan

    2017-09-29

    Guanidinium-functionalized molecules are commonly studied for their use as pharmaceutically active compounds and drugs carriers. Herein, four cyclometalated iridium(III) complexes containing guanidinium ligands have been synthesized and characterized as potential anticancer agents. These complexes exhibit moderate antitumor activity in HeLa, MCF-7, HepG2, CNE-2, and A549 human tumor cells. Interestingly, all complexes showed higher cytotoxicity than cisplatin against a cisplatin-resistant cell line A549R, and less cytotoxicity on the nontumorigenic LO2 cells. Intracellular distribution studies suggest that these complexes are selectively localized in the mitochondria. Mechanism studies indicate that these complexes arrested the cell cycle in the G0/G1 phase and can influence mitochondrial integrity, inducing cancer cell death through reactive oxygen species (ROS)-dependent pathways. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  11. Mitochondria-targeting cyclometalated iridium(III)-PEG complexes with tunable photodynamic activity.

    PubMed

    Li, Steve Po-Yam; Lau, Chris Tsan-Shing; Louie, Man-Wai; Lam, Yun-Wah; Cheng, Shuk Han; Lo, Kenneth Kam-Wing

    2013-10-01

    We present a new class of phosphorescent cyclometalated iridium(III) polypyridine poly(ethylene glycol) (PEG) complexes [Ir(N(^)C)2(bpy-CONH-PEG)](PF6) (bpy-CONH-PEG = 4-(N-(2-(ω-methoxypoly-(1-oxapropyl))ethyl)aminocarbonyl)-4'-methyl-2,2'-bipyridine, number average molecular weight (Mn) = 5272.23, weight average molecular weight (Mw) = 5317.38, polydispersity index (PDI) = 1.009; HN(^)C = 2-phenylpyridine, Hppy (1a), 2-((1,1'-biphenyl)-4-yl)pyridine, Hpppy (2a), 2-phenylquinoline, Hpq (3a), 2-phenylbenzothiazole, Hbt (4a), 2-(1-naphthyl)benzothiazole, Hbsn (5a)). The photophysical, photochemical, and biological properties of these complexes have been compared with those of their PEG-free counterparts [Ir(N(^)C)2(bpy-CONH-Et)](PF6) (bpy-CONH-Et = 4-(N-ethylaminocarbonyl)-4'-methyl-2,2'-bipyridine; HN(^)C = Hppy (1b), Hpppy (2b), Hpq (3b), Hbt (4b), Hbsn (5b)). Upon irradiation, all the complexes exhibited intense and long-lived green to orange-red emission under ambient conditions. The emission was phosphorescence in nature and can be quenched by O2 with the generation of singlet oxygen ((1)O2). The quantum yields for (1)O2 production of the complexes in aerated DMSO (0.24-0.83) were found to be dependent on the excited-state lifetimes of the complexes, which can be altered using different cyclometalating ligands (N(^)C). Cell-based assays indicated that the PEG complexes were noncytotoxic in the dark (IC50 > 300 μM); however, most of them became significantly cytotoxic upon irradiation (IC50 = 3.4 - 23.2 μM). Laser-scanning confocal microscopy images revealed localization of complex 3a in the mitochondrial region of HeLa cells and the induction of rapid necrotic cell death upon light activation. Additionally, the lack of dark toxicity and potential application of the PEG complexes as a visualizing reagent have been demonstrated using zebrafish (Danio rerio) as an animal model. Copyright © 2013 Elsevier Ltd. All rights reserved.

  12. Mitochondria-targeting phosphorescent iridium(III) complexes for living cell imaging.

    PubMed

    Zhang, Qingqing; Cao, Rui; Fei, Hao; Zhou, Ming

    2014-11-28

    Two phosphorescent iridium(III) complexes conjugated to a lipophilic triphenylphosphonium cation moiety, IrMitoOlivine and IrMitoNIR, were synthesized. The complexes show high mitochondria-specificity and relatively lower cytotoxicity. Time-lapse confocal imaging indicates that both complexes exhibit an excellent anti-photobleaching capability under continuous laser irradiation.

  13. Mitochondrial Dysfunction and Oxidative Stress in Asthma: Implications for Mitochondria-Targeted Antioxidant Therapeutics

    PubMed Central

    Reddy, P. Hemachandra

    2011-01-01

    Asthma is a complex, inflammatory disorder characterized by airflow obstruction of variable degrees, bronchial hyper-responsiveness, and airway inflammation. Asthma is caused by environmental factors and a combination of genetic and environmental stimuli. Genetic studies have revealed that multiple loci are involved in the etiology of asthma. Recent cellular, molecular, and animal-model studies have revealed several cellular events that are involved in the progression of asthma, including: increased Th2 cytokines leading to the recruitment of inflammatory cells to the airway, and an increase in the production of reactive oxygen species and mitochondrial dysfunction in the activated inflammatory cells, leading to tissue injury in the bronchial epithelium. Further, aging and animal model studies have revealed that mitochondrial dysfunction and oxidative stress are involved and play a large role in asthma. Recent studies using experimental allergic asthmatic mouse models and peripheral cells and tissues from asthmatic humans have revealed antioxidants as promising treatments for people with asthma. This article summarizes the latest research findings on the involvement of inflammatory changes, and mitochondrial dysfunction/oxidative stress in the development and progression of asthma. This article also addresses the relationship between aging and age-related immunity in triggering asthma, the antioxidant therapeutic strategies in treating people with asthma. PMID:21461182

  14. Neuroprotection and Anti-Epileptogenesis with a Mitochondria-Targeted Antioxidant

    DTIC Science & Technology

    2013-12-01

    3 days after SE, sectioned through the hippocampus and stained for: Nissl , Fluoro-jade C (FJ), NeuN and heat shock protein (HSP). Three rats from...each group died during SE. There was no significant difference in the latency to SE between the two groups. There was also no difference in Nissl , FJ...immunohistochemical stains: Nissl , Fluoro-jade C (FJ), NeuN and heat shock protein 70-72 (HSP). Nissl , FJ and NeuN stains were used to assess

  15. Towards Water Soluble Mitochondria-Targeting Theranostic Osmium(II) Triazole-Based Complexes.

    PubMed

    Omar, Salem A E; Scattergood, Paul A; McKenzie, Luke K; Bryant, Helen E; Weinstein, Julia A; Elliott, Paul I P

    2016-10-18

    The complex [Os(btzpy)₂][PF₆]₂ (1, btzpy = 2,6-bis(1-phenyl-1,2,3-triazol-4-yl)pyridine) has been prepared and characterised. Complex 1 exhibits phosphorescence (λem = 595 nm, τ = 937 ns, φem = 9.3% in degassed acetonitrile) in contrast to its known ruthenium(II) analogue, which is non-emissive at room temperature. The complex undergoes significant oxygen-dependent quenching of emission with a 43-fold reduction in luminescence intensity between degassed and aerated acetonitrile solutions, indicating its potential to act as a singlet oxygen sensitiser. Complex 1 underwent counterion metathesis to yield [Os(btzpy)₂]Cl₂ (1(Cl)), which shows near identical optical absorption and emission spectra to those of 1. Direct measurement of the yield of singlet oxygen sensitised by 1(Cl) was carried out (φ (¹O₂) = 57%) for air equilibrated acetonitrile solutions. On the basis of these photophysical properties, preliminary cellular uptake and luminescence microscopy imaging studies were conducted. Complex 1(Cl) readily entered the cancer cell lines HeLa and U2OS with mitochondrial staining seen and intense emission allowing for imaging at concentrations as low as 1 μM. Long-term toxicity results indicate low toxicity in HeLa cells with LD50 >100 μM. Osmium(II) complexes based on 1 therefore present an excellent platform for the development of novel theranostic agents for anticancer activity.

  16. Triple-responsive Expansile Nanogel for Tumor and Mitochondria Targeted Photosensitizer Delivery

    PubMed Central

    He, Huacheng; Cattran, Alexander W.; Nguyen, Tu; Nieminen, Anna-Liisa

    2014-01-01

    A pH, thermal, and redox potential triple-responsive expansile nanogel system (TRN), which swells at acidic pH, temperature higher than its transition temperature, and reducing environment, has been developed. TRN quickly expands from 108 nm to over 1200 nm (in diameter), achieving more than 1000-fold size enlargement (in volume), within 2 h in a reducing environment at body temperature. Sigma-2 receptor targeting-ligand functionalized TRN can effectively target head and neck tumor, and help Pc 4 targeting mitochondria inside cancer cells to achieve enhanced photodynamic therapy efficacy. PMID:25154666

  17. Intraoral Mitochondrial-Targeted GS-Nitroxide, JP4-039, Radioprotects Normal Tissue in Tumor-Bearing Radiosensitive Fancd2−/− (C57BL/6) Mice

    PubMed Central

    Shinde, Ashwin; Berhane, Hebist; Rhieu, Byung Han; Kalash, Ronny; Xu, Karen; Goff, Julie; Epperly, Michael W.; Franicola, Darcy; Zhang, Xichen; Dixon, Tracy; Shields, Donna; Wang, Hong; Wipf, Peter; Parmar, Kalindi; Guinan, Eva; Kagan, Valerian; Tyurin, Vladimir; Ferris, Robert L.; Zhang, Xiaolan; Li, Song; Greenberger, Joel S.

    2016-01-01

    We evaluated normal tissue specific radioprotection of the oral cavity in radiosensitive Fanconi Anemia (FA) Fancd2−/−mice with orally established tumors using mitochondrial-targeted GS-nitroxide (JP4-039). Adult (10–12 weeks old) Fancd2+/+, Fancd2+/− and Fancd2−/− mice (C57BL/6 background) and subgroups with orally established TC-1 epithelial cell tumors received a single fraction of 28 Gy or four daily fractions of 8 Gy to the head and neck. Subgroups received JP4-039 in F15 emulsion (F15/JP4-039; 0.4 mg/mouse), 4-amino-Tempo in F15 emulsion (F15/4-amino-Tempo; 0.2 mg/ mouse) or F15 emulsion alone prior to each irradiation. Oral mucosa of Fancd2−/− mice showed baseline elevated RNA transcripts for Sod2, p53, p21 and Rad51 (all P < 0.0012) and suppressed levels of Nfkb and Tgfb, (all P < 0.0020) compared with Fancd2+/+ mice. The oral mucosa in tumor-bearing mice of all genotypes showed decreased levels of p53 and elevated Tgfb and Gadd45a (P ≤ 0.0001 for all three genotypes). Intraoral F15/JP4-039, but not F15/4-amino-Tempo, modulated radiation-induced normal tissue transcript elevation, ameliorated mucosal ulceration and reduced the depletion of antioxidant stores in oral cavity tissue of all genotypes, but did not radioprotect tumors. Mitochondrial targeting makes F15/JP4-039 an effective normal tissue radioprotector for Fancd2−/− mice, as well as wild-type mice. PMID:26789701

  18. Electron paramagnetic resonance line shifts and line shape changes due to heisenberg spin exchange and dipole-dipole interactions of nitroxide free radicals in liquids 8. Further experimental and theoretical efforts to separate the effects of the two interactions.

    PubMed

    Peric, Mirna; Bales, Barney L; Peric, Miroslav

    2012-03-22

    The work in part 6 of this series (J. Phys. Chem. A 2009, 113, 4930), addressing the task of separating the effects of Heisenberg spin exchange (HSE) and dipole-dipole interactions (DD) on electron paramagnetic resonance (EPR) spectra of nitroxide spin probes in solution, is extended experimentally and theoretically. Comprehensive measurements of perdeuterated 2,2,6,6-tetramethyl-4-oxopiperidine-1-oxyl (pDT) in squalane, a viscous alkane, paying special attention to lower temperatures and lower concentrations, were carried out in an attempt to focus on DD, the lesser understood of the two interactions. Theoretically, the analysis has been extended to include the recent comprehensive treatment by Salikhov (Appl. Magn. Reson. 2010, 38, 237). In dilute solutions, both interactions (1) introduce a dispersion component, (2) broaden the lines, and (3) shift the lines. DD introduces a dispersion component proportional to the concentration and of opposite sign to that of HSE. Equations relating the EPR spectral parameters to the rate constants due to HSE and DD have been derived. By employing nonlinear least-squares fitting of theoretical spectra to a simple analytical function and the proposed equations, the contributions of the two interactions to items 1-3 may be quantified and compared with the same parameters obtained by fitting experimental spectra. This comparison supports the theory in its broad predictions; however, at low temperatures, the DD contribution to the experimental dispersion amplitude does not increase linearly with concentration. We are unable to deduce whether this discrepancy is due to inadequate analysis of the experimental data or an incomplete theory. A new key aspect of the more comprehensive theory is that there is enough information in the experimental spectra to find items 1-3 due to both interactions; however, in principle, appeal must be made to a model of molecular diffusion to separate the two. The permanent diffusion model is used to

  19. EPR Line Shifts and Line Shape Changes Due to Heisenberg Spin Exchange and Dipole-Dipole Interactions of Nitroxide Free Radicals in Liquids: 8. Further Experimental and Theoretical Efforts to Separate the Effects of the Two Interactions

    PubMed Central

    Peric, Mirna; Bales, Barney L; Peric, Miroslav

    2012-01-01

    The work in Part 6 of this series (J. Phys. Chem. A 2009, 113, 4930), addressing the task of separating the effects of Heisenberg spin exchange (HSE) and dipole-dipole (DD) interactions on EPR spectra of nitroxide spin probes in solution, is extended experimentally and theoretically. Comprehensive measurements of perdeuterated 2,2,6,6-tetramethyl-4-oxopiperidine-1-oxyl (pDT) in squalane, a viscous alkane, paying special attention to lower temperatures and lower concentrations were carried out in an attempt to focus on DD, the lesser understood of the two interactions. Theoretically, the analysis has been extended to include the recent comprehensive treatment by Salikhov (Appl. Magn. Reson. 2010, 38, 237). In dilute solutions, both interactions (1) introduce a dispersion component, (2) broaden the lines, and (3) shift the lines. DD introduces a dispersion component proportional to the concentration and of opposite sign to that of HSE. Equations relating the EPR spectral parameters to the rate constants due HSE and DD have been derived. By employing non-linear least-squares fitting of theoretical spectra to a simple analytical function and the proposed equations, the contributions of the two interactions to items (1)–(3) may be quantified and compared with the same parameters obtained by fitting experimental spectra. This comparison supports the theory in its broad predictions, however, at low temperatures, the DD contribution to the experimental dispersion amplitude does not increase linearly with concentration. We are unable to deduce if this discrepancy is due to inadequate analysis of the experimental data or an incomplete theory. A key new aspect of the more comprehensive theory is that there is enough information in the experimental spectra to find items (1)–(3) due to both interactions; however, in principle, appeal must be made to a model of molecular diffusion to separate the two. The permanent diffusion model is used to illustrate the separation in this

  20. Amelioration of radiation-induced oral cavity mucositis and distant bone marrow suppression in fanconi anemia Fancd2-/- (FVB/N) mice by intraoral GS-nitroxide JP4-039.

    PubMed

    Berhane, Hebist; Shinde, Ashwin; Kalash, Ronny; Xu, Karen; Epperly, Michael W; Goff, Julie; Franicola, Darcy; Zhang, Xichen; Dixon, Tracy; Shields, Donna; Wang, Hong; Wipf, Peter; Li, Song; Gao, Xiang; Greenberger, Joel S

    2014-07-01

    The altered DNA damage response pathway in patients with Fanconi anemia (FA) may increase the toxicity of clinical radiotherapy. We quantitated oral cavity mucositis in irradiated Fanconi anemia Fancd2(-/-) mice, comparing this to Fancd2(+/-) and Fancd2(+/+) mice, and we measured distant bone marrow suppression and quantitated the effect of the intraoral radioprotector GS-nitroxide, JP4-039 in F15 emulsion. We found that FA mice were more susceptible to radiation injury and that protection from radiation injury by JP4-039/F15 was observed at all radiation doses. Adult 10-12-week-old mice, of FVB/N background Fancd2(-/-), Fancd2(+/-) and Fancd2(+/+) were head and neck irradiated with 24, 26, 28 or 30 Gy (large fraction sizes typical of stereotactic radiosurgery treatments) and subgroups received intraoral JP4-039 (0.4 mg/mouse in 100 μL F15 liposome emulsion) preirradiation. On day 2 or 5 postirradiation, mice were sacrificed, tongue tissue and femur marrow were excised for quantitation of radiation-induced stress response, inflammatory and antioxidant gene transcripts, histopathology and assay for femur marrow colony-forming hematopoietic progenitor cells. Fancd2(-/-) mice had a significantly higher percentage of oral mucosal ulceration at day 5 after 26 Gy irradiation (59.4 ± 8.2%) compared to control Fancd2(+/+) mice (21.7 ± 2.9%, P = 0.0063). After 24 Gy irradiation, Fancd2(-/-) mice had a higher oral cavity percentage of tongue ulceration compared to Fancd2(+/+) mice irradiated with higher doses of 26 Gy (P = 0.0123). Baseline and postirradiation oral cavity gene transcripts were altered in Fancd2(-/-) mice compared to Fancd2(+/+) controls. Fancd2(-/-) mice had decreased baseline femur marrow CFU-GM, BFUe and CFU-GEMM, which further decreased after 24 or 26 Gy head and neck irradiation. These changes were not seen in head- and neck-irradiated Fancd2(+/+) mice. In radiosensitive Fancd2(-/-) mice, biomarkers of both local oral cavity and distant marrow

  1. Amelioration of Radiation-Induced Oral Cavity Mucositis and Distant Bone Marrow Suppression in Fanconi Anemia Fancd2−/− (FVB/N) Mice by Intraoral GS-Nitroxide JP4-039

    PubMed Central

    Berhane, Hebist; Shinde, Ashwin; Kalash, Ronny; Xu, Karen; Epperly, Michael W.; Goff, Julie; Franicola, Darcy; Zhang, Xichen; Dixon, Tracy; Shields, Donna; Wang, Hong; Wipf, Peter; Li, Song; Gao, Xiang; Greenberger, Joel S.

    2014-01-01

    The altered DNA damage response pathway in patients with Fanconi anemia (FA) may increase the toxicity of clinical radiotherapy. We quantitated oral cavity mucositis in irradiated Fanconi anemia Fancd2−/− mice, comparing this to Fancd2+/− and Fancd2+/+ mice, and we measured distant bone marrow suppression and quantitated the effect of the intraoral radioprotector GS-nitroxide, JP4-039 in F15 emulsion. We found that FA mice were more susceptible to radiation injury and that protection from radiation injury by JP4-039/F15 was observed at all radiation doses. Adult 10–12-week-old mice, of FVB/N background Fancd2−/−, Fancd2+/− and Fancd2+/+ were head and neck irradiated with 24, 26, 28 or 30 Gy (large fraction sizes typical of stereotactic radiosurgery treatments) and subgroups received intraoral JP4-039 (0.4 mg/mouse in 100 μL F15 liposome emulsion) preirradiation. On day 2 or 5 postirradiation, mice were sacrificed, tongue tissue and femur marrow were excised for quantitation of radiation-induced stress response, inflammatory and antioxidant gene transcripts, histopathology and assay for femur marrow colony-forming hematopoietic progenitor cells. Fancd2−/− mice had a significantly higher percentage of oral mucosal ulceration at day 5 after 26 Gy irradiation (59.4 ± 8.2%) compared to control Fancd2+/+ mice (21.7 ± 2.9%, P = 0.0063). After 24 Gy irradiation, Fancd2−/− mice had a higher oral cavity percentage of tongue ulceration compared to Fancd2+/+ mice irradiated with higher doses of 26 Gy (P = 0.0123). Baseline and postirradiation oral cavity gene transcripts were altered in Fancd2−/− mice compared to Fancd2+/+ controls. Fancd2−/− mice had decreased baseline femur marrow CFU-GM, BFUe and CFU-GEMM, which further decreased after 24 or 26 Gy head and neck irradiation. These changes were not seen in head- and neck-irradiated Fancd2+/+ mice. In radiosensitive Fancd2−/− mice, biomarkers of both local oral cavity and distant marrow

  2. Novel Mitochondria-Targeted Heat-Soluble Proteins Identified in the Anhydrobiotic Tardigrade Improve Osmotic Tolerance of Human Cells

    PubMed Central

    Tanaka, Sae; Tanaka, Junko; Miwa, Yoshihiro; Horikawa, Daiki D.; Katayama, Toshiaki; Arakawa, Kazuharu; Toyoda, Atsushi; Kubo, Takeo; Kunieda, Takekazu

    2015-01-01

    Tardigrades are able to tolerate almost complete dehydration through transition to a metabolically inactive state, called “anhydrobiosis”. Late Embryogenesis Abundant (LEA) proteins are heat-soluble proteins involved in the desiccation tolerance of many anhydrobiotic organisms. Tardigrades, Ramazzottius varieornatus, however, express predominantly tardigrade-unique heat-soluble proteins: CAHS (Cytoplasmic Abundant Heat Soluble) and SAHS (Secretory Abundant Heat Soluble) proteins, which are secreted or localized in most intracellular compartments, except the mitochondria. Although mitochondrial integrity is crucial to ensure cellular survival, protective molecules for mitochondria have remained elusive. Here, we identified two novel mitochondrial heat-soluble proteins, RvLEAM and MAHS (Mitochondrial Abundant Heat Soluble), as potent mitochondrial protectants from Ramazzottius varieornatus. RvLEAM is a group3 LEA protein and immunohistochemistry confirmed its mitochondrial localization in tardigrade cells. MAHS-green fluorescent protein fusion protein localized in human mitochondria and was heat-soluble in vitro, though no sequence similarity with other known proteins was found, and one region was conserved among tardigrades. Furthermore, we demonstrated that RvLEAM protein as well as MAHS protein improved the hyperosmotic tolerance of human cells. The findings of the present study revealed that tardigrade mitochondria contain at least two types of heat-soluble proteins that might have protective roles in water-deficient environments. PMID:25675104

  3. Novel mitochondria-targeted heat-soluble proteins identified in the anhydrobiotic Tardigrade improve osmotic tolerance of human cells.

    PubMed

    Tanaka, Sae; Tanaka, Junko; Miwa, Yoshihiro; Horikawa, Daiki D; Katayama, Toshiaki; Arakawa, Kazuharu; Toyoda, Atsushi; Kubo, Takeo; Kunieda, Takekazu

    2015-01-01

    Tardigrades are able to tolerate almost complete dehydration through transition to a metabolically inactive state, called "anhydrobiosis". Late Embryogenesis Abundant (LEA) proteins are heat-soluble proteins involved in the desiccation tolerance of many anhydrobiotic organisms. Tardigrades, Ramazzottius varieornatus, however, express predominantly tardigrade-unique heat-soluble proteins: CAHS (Cytoplasmic Abundant Heat Soluble) and SAHS (Secretory Abundant Heat Soluble) proteins, which are secreted or localized in most intracellular compartments, except the mitochondria. Although mitochondrial integrity is crucial to ensure cellular survival, protective molecules for mitochondria have remained elusive. Here, we identified two novel mitochondrial heat-soluble proteins, RvLEAM and MAHS (Mitochondrial Abundant Heat Soluble), as potent mitochondrial protectants from Ramazzottius varieornatus. RvLEAM is a group3 LEA protein and immunohistochemistry confirmed its mitochondrial localization in tardigrade cells. MAHS-green fluorescent protein fusion protein localized in human mitochondria and was heat-soluble in vitro, though no sequence similarity with other known proteins was found, and one region was conserved among tardigrades. Furthermore, we demonstrated that RvLEAM protein as well as MAHS protein improved the hyperosmotic tolerance of human cells. The findings of the present study revealed that tardigrade mitochondria contain at least two types of heat-soluble proteins that might have protective roles in water-deficient environments.

  4. Mitochondria-Targeted Photodynamic Therapy with a Galactodendritic Chlorin to Enhance Cell Death in Resistant Bladder Cancer Cells.

    PubMed

    Pereira, Patrícia M R; Silva, Sandrina; Bispo, Mafalda; Zuzarte, Mónica; Gomes, Célia; Girão, Henrique; Cavaleiro, José A S; Ribeiro, Carlos A F; Tomé, João P C; Fernandes, Rosa

    2016-11-16

    Here, we report the rational design of a new third-generation photosensitizer (PS), a chlorin conjugated with galactodendritic units, ChlGal8, to improve the effectiveness of bladder cancer treatment. ChlGal8 shows better photochemical and photophysical properties than a recently reported homologous porphyrin, PorGal8. In addition to inheriting excellent photostability, the ability to generate singlet oxygen, and the ability to interact with the proteins galectin-1 and human serum albumin (HSA), ChlGal8 exhibits high absorption in the red region of the electromagnetic spectrum. In vitro studies of anticancer activity of ChlGal8 revealed that once this PS is taken up by UM-UC-3 bladder cancer cells, it induces high cytotoxicity after a single dose of light irradiation. In HT-1376 bladder cancer cells resistant to therapy, a second light irradiation treatment enhanced in vitro and in vivo photodynamic efficacy. The enhanced phototoxicity in HT-1376 cancer cells seems to be due to the ability of ChlGal8 to accumulate in the mitochondria, via facilitative glucose transporter 1 (GLUT1), in the period between single and repeated irradiation. A photodynamic therapy (PDT) regimen using an extra dose of light irradiation and ChlGal8 as PS represents a promising strategy in treating resistant cancers in a clinical setting.

  5. Structure-Guided Design and Synthesis of a Mitochondria-Targeting Near-Infrared Fluorophore with Multimodal Therapeutic Activities.

    PubMed

    Tan, Xu; Luo, Shenglin; Long, Lei; Wang, Yu; Wang, Dechun; Fang, Shengtao; Ouyang, Qin; Su, Yongping; Cheng, Tianmin; Shi, Chunmeng

    2017-10-05

    An urgent challenge for imaging-guided disease-targeted multimodal therapy is to develop the appropriate multifunctional agents to meet the requirements for potential applications. Here, a rigid cyclohexenyl substitution in the middle of a polymethine linker and two asymmetrical amphipathic N-alkyl side chains to indocyanine green (ICG) (the only FDA-approved NIR contrast agent) are introduced, and a new analog, IR-DBI, is developed with simultaneous cancer-cell mitochondrial targeting, NIR imaging, and chemo-/PDT/PTT/multimodal therapeutic activities. The asymmetrical and amphipathic structural modification renders IR-DBI a close binding to albumin protein site II to form a drug-protein complex and primarily facilitates its preferential accumulation at tumor sites via the enhanced permeability and retention (EPR) effect. The released IR-DBI dye is further actively taken up by cancer cells through organic-anion-transporting polypeptide transporters, and the lipophilic cationic property leads to its selective accumulation in the mitochondria of cancer cells. Finally, based on the high albumin-binding affinity, IR-DBI is modified into human serum albumin (HSA) via self-assembly to produce a nanosized complex, which exhibits significant improvement in the cancer targeting and multimodal cancer treatment with better biocompatibility. This finding may present a practicable strategy to develop small-molecule-based cancer theranostic agents for simultaneous cancer diagnostics and therapeutics. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. [Protective effect of mitochondria-targeted antioxidant SS31 on early brain injury following subarachnoid hemorrhage in rats].

    PubMed

    Zhou, Jian; Li, Zhengzheng; Chen, Zhenggang; Yang, Kun

    2017-09-28

    To evaluate protective effects of SS31 on early brain injury (EBI) induced by subarachnoid hemorrhage (SAH) in rats.
 Methods: A total of 96 Sprague-Dawley rats were randomly divided into 4 groups: A sham group, an SAH group, an SAH+vehicle group (SAH+V), and an SAH+SS31 group. The SAH-induced prechiasmatic cistern rat model was established in this study. Neurological deficit scores were evaluated at 24 h after SAH. The SS31 (5 mg/kg) as well as equal volume of vehicle were administrated intraperitoneally at 2 h after SAH. The neurological scores, brain edema, blood-brain barrier (BBB) permeability, apoptosis, malondialdehyde (MDA), glutathione peroxidase (GPx) activity, superoride dismutase (SOD) activity, and the expression of cytosolic cytochrome c (Cyt C) and Bax were analyzed at 24 h after SAH.
 Results: Treatment with SS31 could significantly reduce MDA levels, and restored the activities of GPx and SOD in the cortex following SAH when compared with the SAH+V group. In addition, Bax SS31 trearment increased or decreased the levels of mitochondrial Cyt C or Bax, respectively. Moreover, SS31 treatment ameliorated brain edema and Evans blue dye extravasation, improved neurological deficits, and decreased neuronal apoptosis at 24 h after SAH.
 Conclusion: SS31 could alleviate EBI after SAH through its antioxidant property and ability in inhibition of neuronal apoptosis.

  7. Mitochondria Targetable Time-Gated Luminescence Probe for Singlet Oxygen Based on a β-Diketonate-Europium Complex.

    PubMed

    Sun, Jingyan; Song, Bo; Ye, Zhiqiang; Yuan, Jingli

    2015-12-21

    Singlet oxygen ((1)O2) plays a key role in the photodynamic therapy (PDT) technique of neoplastic diseases. In this work, by using a 9,10-dimethyl-2-anthryl-containing β-diketone, 1,1,1,2,2-pentafluoro-5-(9',10'-dimethyl-2'-anthryl)-3,5-pentanedione (Hpfdap), as a (1)O2-recognition ligand, a novel β-diketonate-europium(III) complex that can act as a luminescence probe for (1)O2, [Eu(pfdap)3(tpy)] (tpy = 2,2',2″-terpyridine), has been designed and synthesized for the time-gated luminescence detection of (1)O2 in living cells. The complex is weakly luminescent due to the quenching effect of 9,10-dimethyl-2-anthryl groups. After reaction with (1)O2, accompanied by the formation of endoperoxides of 9,10-dimethyl-2-anthryl groups, the luminescence quenching disappears, so that the long-lived luminescence of the europium(III) complex is switched on. The complex showed highly selective luminescence response to (1)O2 with a remarkable luminescence enhancement. Combined with the time-gated luminescence imaging technique, the complex was successfully used as a luminescent probe for the monitoring of the time-dependent generation of (1)O2 in 5-aminolevulinic acid (a PDT drug) loaded HepG2 cells during the photodynamic process. In addition, by coloading the complex and a mitochondrial indicator, Mito-Tracker Green, into HepG2 cells, the specific localization of [Eu(pfdap)3(tpy)] molecules in mitochondria of HepG2 cells was demonstrated by confocal fluorescence imaging measurements.

  8. Cancer chemoprevention and mitochondria: targeting apoptosis in transformed cells via the disruption of mitochondrial bioenergetics/redox state.

    PubMed

    Hail, Numsen; Lotan, Reuben

    2009-01-01

    Cancer chemoprevention employs agents that block, hinder, or reverse tumorigenesis to prevent malignancy. Several putative cancer chemopreventive agents promote apoptosis in transformed cells initiated in animal carcinogenesis models or identified in human subjects, and/or in tumor cells cultured in vitro. Consequently, apoptosis induction is increasingly valued as a biologically significant anticancer mechanism in the arena of chemoprevention. In vitro studies suggest that the permeabilization of mitochondrial membranes is an important mechanistic determinant associated with the apoptosis induced by these agents. Mitochondrial membrane permeabilization (MMP) may occur via the control of proapoptotic Bcl-2 family members, and/or by the induction of the mitochondrial permeability transition. Both of these cell death-inducing regulatory mechanisms are ultimately responsive to the bioenergetic status/redox state of mitochondria. Interestingly, in addition to inducing MMP, various chemopreventive agents can directly modulate mitochondrial bioenergetics and/or redox tone in transformed cells. This review will examine prospective mechanisms associated with the disruption of mitochondrial function by chemopreventive agents that affect MMP and apoptosis. In doing so, we will construct a paradigm supporting the notion that the bioenergetic and/or redox characteristics of the mitochondria in transformed cells are important targets in the chemoprevention of cancer.

  9. Integrating Enzymatic Self-Assembly and Mitochondria Targeting for Selectively Killing Cancer Cells without Acquired Drug Resistance.

    PubMed

    Wang, Huaimin; Feng, Zhaoqianqi; Wang, Youzhi; Zhou, Rong; Yang, Zhimou; Xu, Bing

    2016-12-14

    Targeting organelles by modulating the redox potential of mitochondria is a promising approach to kill cancer cells that minimizes acquired drug resistance. However, it lacks selectivity because mitochondria perform essential functions for (almost) all cells. We show that enzyme-instructed self-assembly (EISA), a bioinspired molecular process, selectively generates the assemblies of redox modulators (e.g., triphenyl phosphinium (TPP)) in the pericellular space of cancer cells for uptake, which allows selectively targeting the mitochondria of cancer cells. The attachment of TPP to a pair of enantiomeric, phosphorylated tetrapeptides produces the precursors (L-1P or D-1P) that form oligomers. Upon dephosphorylation catalyzed by ectophosphatases (e.g., alkaline phosphatase (ALP)) overexpressed on cancer cells (e.g., Saos2), the oligomers self-assemble to form nanoscale assemblies only on the surface of the cancer cells. The cancer cells thus uptake these assemblies of TPP via endocytosis, mainly via a caveolae/raft-dependent pathway. Inside the cells, the assemblies of TPP-peptide conjugates escape from the lysosome, induce dysfunction of mitochondria to release cytochrome c, and result in cell death, while the controls (i.e., omitting TPP motif, inhibiting ALP, or removing phosphate trigger) hardly kill the Saos2 cells. Most importantly, the repeated stimulation of the cancers by the precursors, unexpectedly, sensitizes the cancer cells to the precursors. As the first example of the integration of subcellular targeting with cell targeting, this study validates the spatial control of the assemblies of nonspecific cytotoxic agents by EISA as a promising molecular process for selectively killing cancer cells without inducing acquired drug resistance.

  10. Highly selective mitochondria-targeting amphiphilic silicon(IV) phthalocyanines with axially ligated rhodamine B for photodynamic therapy.

    PubMed

    Zhao, Zhixin; Chan, Pui-Shan; Li, Hongguang; Wong, Ka-Leung; Wong, Ricky Ngok Shun; Mak, Nai-Ki; Zhang, Jie; Tam, Hoi-Lam; Wong, Wai-Yeung; Kwong, Daniel W J; Wong, Wai-Kwok

    2012-01-16

    Two axially ligated rhodamine-Si(IV)-phthalocyanine (Rh-SiPc) conjugates, bearing one and two rhodamine B, were synthesized and their linear and two-photon photophysical, subcellular localization and photocytotoxic properties were studied. These Rh-SiPc conjugates exhibited an almost exclusive mitochondrial localizing property in human nasopharyngeal carcinoma (HK-1) cells and human cervical carcinoma (HeLa) cells. Strong photocytotoxic but low dark cytotoxic properties were also observed for the two Rh-SiPc conjugates toward the HK-1 cells. Using nuclei staining method and flow cytometric DNA content analysis, apoptotic cell death was induced by these conjugates upon photoactivation. This observation is consistent with their mitochondrial localization property. The observed properties of these conjugates qualify them as promising PDT agents.

  11. Proteome-scale identification and characterization of mitochondria targeting proteins of Mycobacterium avium subspecies paratuberculosis: Potential virulence factors modulating host mitochondrial function.

    PubMed

    Rana, Aarti; Kumar, Devender; Rub, Abdur; Akhter, Yusuf

    2015-07-01

    Mycobacterium avium subsp. paratuberculosis is the etiological agent of Johne's Disease among ruminants. During the course of infection, it expresses a number of proteins for its successful persistence inside the host that cause variety of physiological abnormalities in the host. Mitochondrion is one of the attractive targets for pathogenic bacteria. Employing a proteome-wide sequence and structural signature based approach we have identified 46 M. avium subsp. paratuberculosis proteins as potential targets for the host mitochondrial targeting. These may act as virulence factors modulating mitochondrial physiology for bacterial survival and immune evasion inside the host cells.

  12. A double-blind, placebo-controlled study to assess the mitochondria-targeted antioxidant MitoQ as a disease-modifying therapy in Parkinson's disease.

    PubMed

    Snow, Barry J; Rolfe, Fiona L; Lockhart, Michelle M; Frampton, Christopher M; O'Sullivan, John D; Fung, Victor; Smith, Robin A J; Murphy, Michael P; Taylor, Kenneth M

    2010-08-15

    Multiple lines of evidence point to mitochondrial oxidative stress as a potential pathogenic cause for Parkinson's disease (PD). MitoQ is a powerful mitochondrial antioxidant. It is absorbed orally and concentrates within mitochondria where it has been shown to protect against oxidative damage. We enrolled 128 newly diagnosed untreated patients with PD in a double-blind study of two doses of MitoQ compared with placebo to explore the hypothesis that, over 12 months, MitoQ would slow the progression of PD as measured by clinical scores, particularly the Unified Parkinson Disease Rating Scale. We showed no difference between MitoQ and placebo on any measure of PD progression. MitoQ does not slow the progression of PD, and this finding should be taken into account when considering the oxidative stress hypothesis for the pathogenesis of PD.

  13. Synthesis of 5α,8α-Ergosterol Peroxide 3-Carbamate Derivatives and a Fluorescent Mitochondria-Targeting Conjugate for Enhanced Anticancer Activities.

    PubMed

    Bu, Ming; Cao, Tingting; Li, Hongxia; Guo, Mingzhou; Yang, Burton B; Zeng, Chengchu; Hu, Liming

    2017-03-17

    Inspired by the significant anticancer activity of our previously screened natural ergosterol peroxide (1), we synthesized and characterized a series of novel ergosterol peroxide 3-carbamate derivatives. The antiproliferative activities of the synthesized compounds against human hepatocellular carcinoma cells (HepG2, SK-Hep1) and human breast cancer cells (MCF-7, MDA-MB231) were investigated. 5α,8α-Epidioxyergosta-3-yl-(piperazine-1)carbamate (3 d) and 5α,8α-epidioxyergosta-3-yl-(piperidin-4-methylamine)carbamate (3 f) and their hydrochloride salts exhibited significant in vitro antiproliferative activities against the tested tumor cell lines, with IC50 values ranging from 0.85 to 4.62 μm. Furthermore, fluorescent imaging showed that the designed coumarin-3 d conjugate (5) localized mainly in mitochondria, leading to enhanced anticancer activities over the parent structure 1. As a whole, it appeared that substituent changes at the C3 position could serve as a promising launch point for further design of this type of steroidal anticancer agent. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Inhibition of ROS production through mitochondria-targeted antioxidant and mitochondrial uncoupling increases post-thaw sperm viability in yellow catfish.

    PubMed

    Fang, Lu; Bai, Chenglian; Chen, Yuanhong; Dai, Jun; Xiang, Yang; Ji, Xiaoping; Huang, Changjiang; Dong, Qiaoxiang

    2014-12-01

    Reactive oxygen species (ROS) are one of the main causes for decreased viability in cryopreserved sperm. Many studies have reported the beneficial effect of antioxidant supplements in freezing media for post-thaw sperm quality. In the present study, we explored two new approaches of ROS inhibition in sperm cryopreservation of yellow catfish, namely mitochondrial-targeted antioxidant and metabolic modulator targeting mitochondrial uncoupling pathways. Our study revealed that addition of MitoQ, a compound designed to deliver ubiquinone into mitochondria, significantly decreased ROS production, as well as lipid peroxidation, and increased post-thaw viability. Similarly, sperm incubated with 2,4-dinitrophenol (DNP), a chemical protonophore that induces mitochondrial uncoupling, also had reduced ROS production, as well as lipid peroxidation, and increased post-thaw sperm viability. Conversely, activation of uncoupling protein (UCP2) by 4-hydroxynonenal (HNE) neither reduced ROS production nor increased post-thaw sperm viability. Our findings indicate that ROS inhibition through mitochondrial-targeted antioxidant or mild mitochondrial uncoupling is beneficial for sperm cryopreservation in yellow catfish. Our study provides novel methods to mitigate oxidative stress induced damage in cryopreserved sperm for future applications.

  15. Overcoming drug-resistant lung cancer by paclitaxel loaded dual-functional liposomes with mitochondria targeting and pH-response.

    PubMed

    Jiang, Lei; Li, Li; He, Xiaodan; Yi, Qiangying; He, Bin; Cao, Jun; Pan, Weisan; Gu, Zhongwei

    2015-06-01

    Mitochondrion-orientated transportation of smart liposomes has been developed as a promising strategy to deliver anticancer drugs directly to tumor sites, and these have a tremendous potential for killing cancer cells, especially those with multidrug resistance (MDR). Herein we report a novel dual-functional liposome system possessing both extracellular pH response and mitochondrial targeting properties to enhance drug accumulation in mitochondria and trigger apoptosis of drug-resistant cancer cells. Briefly, peptide D[KLAKLAK]2 (KLA) was modified with 2, 3-dimethylmaleic anhydride (DMA) and combined with 1, 2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE) to yield a DSPE-KLA-DMA (DKD) lipid. This dual-functional DKD was then mixed with other commercially available lipids to fabricate liposomes. In vitro anticancer efficacy of this liposome system was evaluated in human lung cancer A549 cells and drug-resistant lung cancer A549/Taxol cells. At tumor extracellular pH (∼6.8), liposomes could reverse their surface charge (negative to positive), facilitating liposome internalization. After cellular uptake, KLA peptide directed delivery-enabled selective accumulation of these liposomes into mitochondria and favored release of their cargo paclitaxel (PTX) into desired sites. Specifically, enhanced apoptosis of MDR cancer cells through mitochondrial signaling pathways was evidenced by release of cytochrome c and increased activity of caspase-9 and -3. These dual-functional liposomes had the greatest efficacy for treating A549 cells and A549/Taxol cells in vitro, and in treating drug-resistant lung cancer A549/Taxol cells xenografted onto nude mice (tumor growth inhibition 86.7%). In conclusion, dual-functional liposomes provide a novel and versatile approach for overcoming MDR in cancer treatment.

  16. Phospho-sulindac (OXT-328) Inhibits the Growth of Human Lung Cancer Xenografts in Mice: Enhanced Efficacy and Mitochondria Targeting by Its Formulation in Solid Lipid Nanoparticles

    PubMed Central

    Zhu, Rongrong; Cheng, Ka-Wing; Mackenzie, Gerardo; Huang, Liqun; Sun, Yu; Xie, Gang; Vrankova, Kveta; Rigas, Basil; Constantinides, Panayiotis P.

    2013-01-01

    Purpose To evaluate the antitumor efficacy of solid lipid nanoparticle–encapsulated phospho-sulindac (SLN-PS) in human lung cancer. Methods PS was incorporated into SLNs using the emulsion evaporation technique. We determined the antitumor activity of SLN-PS in cultured lung cancer cells. The performance of SLN-PS was further evaluated by pharmacokinetic studies in mice and in a model of human lung cancer xenografts in nude mice. Results SLN-PS was >4-fold more potent than PS in inhibiting the growth of A549 and H510 cells in vitro. SLN-PS enhanced cellular uptake and facilitated PS accumulation in mitochondria, leading to oxidative stress and apoptosis via the mitochondrial-apoptosis pathway. SLN-PS was highly effective in suppressing the growth of A549 xenografts (78% inhibition compared to control, p < 0.01); while PS had no significant effect. Formulation of PS in SLNs resulted in improved pharmacokinetics in mice and an enhanced (~14-fold) accumulation of PS and its metabolites in A549 xenografts. Finally, SLN-PS enhanced urinary F2-isoprostane uniquely in mice bearing A549 xenografts compared to untreated controls, suggesting that SLN-PS specifically induced oxidative stress in tumors. Conclusions Our results show that SLN-PS is efficacious in suppressing the growth of lung cancer and merits further evaluation. PMID:22723123

  17. The immature fiber mutant phenotype of cotton (Gossypium hirsutum) is linked to a 22-bp frame-shift deletion in a mitochondria targeted pentatricopeptide repeat gene

    USDA-ARS?s Scientific Manuscript database

    Cotton seed trichomes are the globally most important source of natural fibers. The major fiber thickness properties influence the price of the raw material and the quality of the finished product. The recessive immature fiber (im) gene reduces the degree of fiber cell wall thickening by a process...

  18. The Immature Fiber Mutant Phenotype of Cotton (Gossypium hirsutum) Is Linked to a 22-bp Frame-Shift Deletion in a Mitochondria Targeted Pentatricopeptide Repeat Gene

    PubMed Central

    Thyssen, Gregory N.; Fang, David D.; Zeng, Linghe; Song, Xianliang; Delhom, Christopher D.; Condon, Tracy L.; Li, Ping; Kim, Hee Jin

    2016-01-01

    Cotton seed trichomes are the most important source of natural fibers globally. The major fiber thickness properties influence the price of the raw material, and the quality of the finished product. The recessive immature fiber (im) gene reduces the degree of fiber cell wall thickening by a process that was previously shown to involve mitochondrial function in allotetraploid Gossypium hirsutum. Here, we present the fine genetic mapping of the im locus, gene expression analysis of annotated proteins near the locus, and association analysis of the linked markers. Mapping-by-sequencing identified a 22-bp deletion in a pentatricopeptide repeat (PPR) gene that is completely linked to the immature fiber phenotype in 2837 F2 plants, and is absent from all 163 cultivated varieties tested, although other closely linked marker polymorphisms are prevalent in the diversity panel. This frame-shift mutation results in a transcript with two long open reading frames: one containing the N-terminal transit peptide that targets mitochondria, the other containing only the RNA-binding PPR domains, suggesting that a functional PPR protein cannot be targeted to mitochondria in the im mutant. Taken together, these results suggest that PPR gene Gh_A03G0489 is involved in the cotton fiber wall thickening process, and is a promising candidate gene at the im locus. Our findings expand our understanding of the molecular mechanisms that modulate cotton fiber fineness and maturity, and may facilitate the development of cotton varieties with superior fiber attributes. PMID:27172184

  19. The novel mitochondria-targeted antioxidant SkQ1 modulates angiogenesis and inflammatory micromilieu in a murine orthotopic model of pancreatic cancer.

    PubMed

    Bazhin, Alexandr V; Yang, Yuhui; D'Haese, Jan G; Werner, Jens; Philippov, Pavel P; Karakhanova, Svetlana

    2016-07-01

    Our understanding in the last few years about reactive oxygen species (ROS) has changed from being harmful substances to crucial intra- and extracellular messengers as well as important regulators controlling a wide spectrum of signaling pathways, including those in cancer immunology. Therefore, these multiple essential roles of ROS and especially of mitochondria-derived ROS in malignant transformation and cancer progression make them a promising target for anticancer therapy. Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers in the world. A link between ROS, antioxidants and the PDAC development and progression has been recently established. Therefore, usage of specific highly efficient antioxidants could bring an option for treatment and/or prevention of PDAC. 10-(6'-plastoquinonyl) decyltriphenylphosphonium (SkQ1) is a new antioxidant with the highest mitochondrion membrane penetrating ability and potent antioxidant capability. In this work, we investigated an impact of SkQ1 on tumor angiogenesis, immune micromilieu, and oncological parameters in the orthotopic Panc02 murine model of PDAC. We showed that in this model SkQ1 treatment leads to the elevation of pro-angiogenic factors and to building of mainly an anti-inflammatory cytokine milieu. On the cellular level we showed an increase in a percentage of memory T cells and a decrease in frequency on natural killer T (NKT) cells. At the same time, SkQ1 was ineffective in the improvement of oncological parameters of PDAC-bearing mice. New studies are needed to clarify the absence of therapeutic and/or prophylactic benefits of the antioxidant. © 2016 UICC.

  20. Phospho-sulindac (OXT-328) inhibits the growth of human lung cancer xenografts in mice: enhanced efficacy and mitochondria targeting by its formulation in solid lipid nanoparticles.

    PubMed

    Zhu, Rongrong; Cheng, Ka-Wing; Mackenzie, Gerardo; Huang, Liqun; Sun, Yu; Xie, Gang; Vrankova, Kveta; Constantinides, Panayiotis P; Rigas, Basil

    2012-11-01

    To evaluate the antitumor efficacy of solid lipid nanoparticle-encapsulated phospho-sulindac (SLN-PS) in human lung cancer. PS was incorporated into SLNs using the emulsion evaporation technique. We determined the antitumor activity of SLN-PS in cultured lung cancer cells. The performance of SLN-PS was further evaluated by pharmacokinetic studies in mice and in a model of human lung cancer xenografts in nude mice. SLN-PS was >4-fold more potent than PS in inhibiting the growth of A549 and H510 cells in vitro. SLN-PS enhanced cellular uptake and facilitated PS accumulation in mitochondria, leading to oxidative stress and apoptosis via the mitochondrial-apoptosis pathway. SLN-PS was highly effective in suppressing the growth of A549 xenografts (78% inhibition compared to control, p < 0.01); while PS had no significant effect. Formulation of PS in SLNs resulted in improved pharmacokinetics in mice and an enhanced (≈ 14-fold) accumulation of PS and its metabolites in A549 xenografts. Finally, SLN-PS enhanced urinary F2-isoprostane uniquely in mice bearing A549 xenografts compared to untreated controls, suggesting that SLN-PS specifically induced oxidative stress in tumors. Our results show that SLN-PS is efficacious in suppressing the growth of lung cancer and merits further evaluation.

  1. Hexahalorhenate(iv) salts of metal oxazolidine nitroxides.

    PubMed

    Pedersen, Anders H; Geoghegan, Blaise L; Nichol, Gary S; Lupton, David W; Murray, Keith S; Martínez-Lillo, José; Gass, Ian A; Brechin, Euan K

    2017-04-04

    Eight coordination compounds of formulae [Fe(II)(L˙)2][Re(IV)Cl6] (1a), [Fe(II)(L˙)2][Re(IV)Br6] (1b), [Co(II)(L˙)2][Re(IV)Cl6]·CH3CN (2a), [Co(II)(L˙)2][Re(IV)Br6] (2b), [Ni(II)(L˙)(CH3CN)3][Re(IV)Cl6]·CH3CN (3a), [Ni(II)(L˙)(CH3CN)3][Re(IV)Br6]·3CH3CN (3b), [Cu(II)(L˙)2][Re(IV)Cl6] (4a) and [Cu(II)(L˙)2][Re(IV)Br6] (4b), where L˙ is the aminoxyl radical chelating ligand, 4,4'-dimethyl-2,2'-di(2-pyridyl)oxazolidine-N-oxide, have been synthesised. Structural and magnetic studies reveal metal-radical intramolecular antiferromagnetic interactions in the [M(II)(L˙)2](2+) cations in the iron, cobalt and copper based compounds (1a, 1b, 2a, 2b, 4a and 4b) with the central metal ion low-spin in the case of iron (1a and 1b) and a gradual, cobalt based, spin-crossover transition present in 2a and 2b. The nickel based compounds, 3a and 3b, were analysed in the dried form (3a(dried) and 3b(dried)) and directly in acetonitrile (3a(solvated) and 3b(solvated)). Microanalysis and IR spectroscopy on 3a(dried) and 3b(dried) suggest that the dried samples are best formulated as [Ni(II)(L˙)(H2O)3][Re(IV)X6], where X = Cl (3a(dried)) and Br (3b(dried)). All forms of 3a and 3b exhibit cationic metal-radical ferromagnetic interactions resulting in S = 3/2 ground states. In addition, 3a(dried) exhibits spin-canting behaviour with an ordering temperature of 2.7 K, an open hysteresis loop with a coercive field Hc = 580 Oe, and a remanent magnetisation Mr = 0.21μB, resulting in a canting angle of ∼1.8°. In contrast, 3b(dried) shows no spin-canting behaviour; a maximum in χMvs. T at T = 3 K suggesting long-range antiferromagnetic ordering. 3a(solvated) and 3b(solvated) show no indication of long-range magnetic ordering, unlike 4a and 4b where anomalies are evident in the low-temperature magnetic susceptibility measurements.

  2. Rapid-scan EPR of nitroxide spin labels and semiquinones

    PubMed Central

    Eaton, Sandra S.; Eaton, Gareth R.

    2017-01-01

    Rapid-scan electron paramagnetic resonance is based on continuous direct detection of the spin response as the magnetic field is scanned up-field and down-field through resonance thousands of times per second. The method provides improved signal-to-noise for a wide range of samples, including rapidly tumbling and immobilized radicals. This chapter provides an introduction to the method and practical examples of implementation for organic radicals. PMID:26478479

  3. Novel Nitroxide Resuscitation Strategies in Experimental Traumatic Brain Injury

    DTIC Science & Technology

    2010-03-01

    peroxidase activity after traumatic brain injury. J Neurotrauma 2003;20:437–445. 68. Mavelli I, Rigo A, Federico R, et al. Superoxide dismutase, glu... Engel , C.C., and Castro, C.A. (2008). Mild traumatic brain injury in U.S. soldiers returning from Iraq. N. Engl. J. Med. 358, 453–463. Ling, G., Bandak

  4. A review of the basics of mitochondrial bioenergetics, metabolism, and related signaling pathways in cancer cells: Therapeutic targeting of tumor mitochondria with lipophilic cationic compounds.

    PubMed

    Kalyanaraman, Balaraman; Cheng, Gang; Hardy, Micael; Ouari, Olivier; Lopez, Marcos; Joseph, Joy; Zielonka, Jacek; Dwinell, Michael B

    2017-09-29

    The present review is a sequel to the previous review on cancer metabolism published in this journal. This review focuses on the selective antiproliferative and cytotoxic effects of mitochondria-targeted therapeutics (MTTs) in cancer cells. Emerging research reveals a key role of mitochondrial respiration on tumor proliferation. Previously, a mitochondria-targeted nitroxide was shown to selectively inhibit colon cancer cell proliferation at submicromolar levels. This review is centered on the therapeutic use of MTTs and their bioenergetic profiling in cancer cells. Triphenylphosphonium cation conjugated to a parent molecule (e.g., vitamin-E or chromanol, ubiquinone, and metformin) via a linker alkyl chain is considered an MTT. MTTs selectively and potently inhibit proliferation of cancer cells and, in some cases, induce cytotoxicity. MTTs inhibit mitochondrial complex I activity and induce mitochondrial stress in cancer cells through generation of reactive oxygen species. MTTs in combination with glycolytic inhibitors synergistically inhibit tumor cell proliferation. This review discusses how signaling molecules traditionally linked to tumor cell proliferation affect tumor metabolism and bioenergetics (glycolysis, TCA cycle, and glutaminolysis). Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  5. AP39, a Mitochondria-Targeted Hydrogen Sulfide Donor, Supports Cellular Bioenergetics and Protects against Alzheimer's Disease by Preserving Mitochondrial Function in APP/PS1 Mice and Neurons

    PubMed Central

    Zhao, Feng-li; Fang, Fang; Qiao, Pei-feng; Yan, Ning; Gao, Dan; Yan, Yong

    2016-01-01

    Increasing evidence suggests that mitochondrial functions are altered in AD and play an important role in AD pathogenesis. It has been established that H2S homeostasis is balanced in AD. The emerging mitochondrial roles of H2S include antioxidation, antiapoptosis, and the modulation of cellular bioenergetics. Here, using primary neurons from the well-characterized APP/PS1 transgenic mouse model, we studied the effects of AP39 (a newly synthesized mitochondrially targeted H2S donor) on mitochondrial function. AP39 increased intracellular H2S levels, mainly in mitochondrial regions. AP39 exerted dose-dependent effects on mitochondrial activity in APP/PS1 neurons, including increased cellular bioenergy metabolism and cell viability at low concentrations (25–100 nM) and decreased energy production and cell viability at a high concentration (250 nM). Furthermore, AP39 (100 nM) increased ATP levels, protected mitochondrial DNA, and decreased ROS generation. AP39 regulated mitochondrial dynamics, shifting from fission toward fusion. After 6 weeks, AP39 administration to APP/PS1 mice significantly ameliorated their spatial memory deficits in the Morris water maze and NORT and reduced Aβ deposition in their brains. Additionally, AP39 inhibited brain atrophy in APP/PS1 mice. Based on these results, AP39 was proposed as a promising drug candidate for AD treatment, and its anti-AD mechanism may involve protection against mitochondrial damage. PMID:27057285

  6. AP39, a Mitochondria-Targeted Hydrogen Sulfide Donor, Supports Cellular Bioenergetics and Protects against Alzheimer's Disease by Preserving Mitochondrial Function in APP/PS1 Mice and Neurons.

    PubMed

    Zhao, Feng-Li; Fang, Fang; Qiao, Pei-feng; Yan, Ning; Gao, Dan; Yan, Yong

    2016-01-01

    Increasing evidence suggests that mitochondrial functions are altered in AD and play an important role in AD pathogenesis. It has been established that H2S homeostasis is balanced in AD. The emerging mitochondrial roles of H2S include antioxidation, antiapoptosis, and the modulation of cellular bioenergetics. Here, using primary neurons from the well-characterized APP/PS1 transgenic mouse model, we studied the effects of AP39 (a newly synthesized mitochondrially targeted H2S donor) on mitochondrial function. AP39 increased intracellular H2S levels, mainly in mitochondrial regions. AP39 exerted dose-dependent effects on mitochondrial activity in APP/PS1 neurons, including increased cellular bioenergy metabolism and cell viability at low concentrations (25-100 nM) and decreased energy production and cell viability at a high concentration (250 nM). Furthermore, AP39 (100 nM) increased ATP levels, protected mitochondrial DNA, and decreased ROS generation. AP39 regulated mitochondrial dynamics, shifting from fission toward fusion. After 6 weeks, AP39 administration to APP/PS1 mice significantly ameliorated their spatial memory deficits in the Morris water maze and NORT and reduced Aβ deposition in their brains. Additionally, AP39 inhibited brain atrophy in APP/PS1 mice. Based on these results, AP39 was proposed as a promising drug candidate for AD treatment, and its anti-AD mechanism may involve protection against mitochondrial damage.

  7. Imaging of mitochondrial Ca2+ dynamics in astrocytes using cell-specific mitochondria-targeted GCaMP5G/6s: mitochondrial Ca2+ uptake and cytosolic Ca2+ availability via the endoplasmic reticulum store.

    PubMed

    Li, Hailong; Wang, Xiaowan; Zhang, Nannan; Gottipati, Manoj K; Parpura, Vladimir; Ding, Shinghua

    2014-12-01

    Mitochondrial Ca(2+) plays a critical physiological role in cellular energy metabolism and signaling, and its overload contributes to various pathological conditions including neuronal apoptotic death in neurological diseases. Live cell mitochondrial Ca(2+) imaging is an important approach to understand mitochondrial Ca(2+) dynamics. Recently developed GCaMP genetically-encoded Ca(2+) indicators provide unique opportunity for high sensitivity/resolution and cell type-specific mitochondrial Ca(2+) imaging. In the current study, we implemented cell-specific mitochondrial targeting of GCaMP5G/6s (mito-GCaMP5G/6s) and used two-photon microscopy to image astrocytic and neuronal mitochondrial Ca(2+) dynamics in culture, revealing Ca(2+) uptake mechanism by these organelles in response to cell stimulation. Using these mitochondrial Ca(2+) indicators, our results show that mitochondrial Ca(2+) uptake in individual mitochondria in cultured astrocytes and neurons can be seen after stimulations by ATP and glutamate, respectively. We further studied the dependence of mitochondrial Ca(2+) dynamics on cytosolic Ca(2+) changes following ATP stimulation in cultured astrocytes by simultaneously imaging mitochondrial and cytosolic Ca(2+) increase using mito-GCaMP5G and a synthetic organic Ca(2+) indicator, x-Rhod-1, respectively. Combined with molecular intervention in Ca(2+) signaling pathway, our results demonstrated that the mitochondrial Ca(2+) uptake is tightly coupled with inositol 1,4,5-trisphosphate receptor-mediated Ca(2+) release from the endoplasmic reticulum and the activation of G protein-coupled receptors. The current study provides a novel approach to image mitochondrial Ca(2+) dynamics as well as Ca(2+) interplay between the endoplasmic reticulum and mitochondria, which is relevant for neuronal and astrocytic functions in health and disease.

  8. Micellar cathodes from self-assembled nitroxide-containing block copolymers in battery electrolytes.

    PubMed

    Hauffman, Guillaume; Maguin, Quentin; Bourgeois, Jean-Pierre; Vlad, Alexandru; Gohy, Jean-François

    2014-01-01

    This contribution describes the synthesis of block copolymers containing electrochemically active blocks, their micellization, and finally their use as micellar cathodes in a lithium battery. The self-assembly of the synthesized poly(styrene)-block-poly(2,2,6,6-tetramethylpiperidinyloxy-4-yl methacrylate) (PS-b-PTMA) diblock copolymers is realized in a typical battery electrolyte made of 1 m lithium trifluoromethanesulfonate dissolved in a mixture of ethylene carbonate/diethyl carbonate/dimethyl carbonate(1:1:1, in volume). Dynamic light scattering and atomic force micro-scopy indicate the formation of well-defined spherical micelles with a PS core and a PTMA corona. The electrochemical properties of those micelles are further investigated. Cyclic voltammograms show a reversible redox reaction at 3.6 V (vs Li(+) /Li). The charge/discharge profiles indicate a flat and reversible plateau around 3.6 V (vs Li(+) /Li). Finally, the cycling performances of the micellar cathodes are demonstrated. Such self-assembled block copolymers open new opportunities for nanostructured organic radical batteries. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Magnetic exchange interaction in gadolinium(III) complex having aliphatic nitroxide radical TEMPO

    SciTech Connect

    Nakamura, Takeshi; Ishida, Takayuki

    2016-02-01

    We synthesized a new compound, [Gd(hfac){sub 3}(MeOH)(TEMPO)] (TEMPO = 2,2,6,6-tetramethylpiperidin-1-oxyl; Hhfac = 1,1,1,5,5,5-hexafluoropentane-2,4-dione) with the metal/radical ratio of 1/1. This compound has an advantage in the magnetic analysis, because the exchange coupling system is described with a unique coupling parameter J, when compared to the structure and magnetic properties of the previous TEMPO and related complexes. The X-ray crystal structure analysis of [Gd(hfac){sub 3}(MeOH)(TEMPO)] revealed the N-O and Gd-O(N) bond lengths are 1.299(9) and 2.307(5) Å, respectively, and the Gd-O-N angle is 149.4(5)°. The magnetic study clarified the Gd{sup 3+}-radical antiferromagnetic interaction of 2J/k{sub B} = −3.5(1) K.

  10. Rapid Dihydrogen Cleavage by Persistent Nitroxide Radicals under Frustrated Lewis Pair Conditions.

    PubMed

    Tao, Xin; Kehr, Gerald; Wang, Xiaowu; Daniliuc, Constantin G; Grimme, Stefan; Erker, Gerhard

    2016-07-04

    Persistent radicals undergo hydrogen atom abstraction reactions with a great variety of substrates, but not with dihydrogen. It has now been found that the TEMPO radical splits dihydrogen under mild conditions in the presence of the strong bulky B(C6 F5 )3 boron Lewis acid. The reaction is thought to proceed by a typical frustrated Lewis pair mechanism with the TEMPO radical acting as the active Lewis base. The reaction was analyzed by DFT, which indicates that no significant spin density on the hydrogen atoms is accumulated along the H2 splitting reaction path.

  11. Revealing the Adsorption Mechanisms of Nitroxides on Ultrapure, Metallicity-Sorted Carbon Nanotubes

    PubMed Central

    2014-01-01

    Carbon nanotubes are a natural choice as gas sensor components given their high surface to volume ratio, electronic properties, and capability to mediate chemical reactions. However, a realistic assessment of the interaction of the tube wall and the adsorption processes during gas phase reactions has always been elusive. Making use of ultraclean single-walled carbon nanotubes, we have followed the adsorption kinetics of NO2 and found a physisorption mechanism. Additionally, the adsorption reaction directly depends on the metallic character of the samples. Franck–Condon satellites, hitherto undetected in nanotube–NOx systems, were resolved in the N 1s X-ray absorption signal, revealing a weak chemisorption, which is intrinsically related to NO dimer molecules. This has allowed us to identify that an additional signal observed in the higher binding energy region of the core level C 1s photoemission signal is due to the C=O species of ketene groups formed as reaction byproducts . This has been supported by density functional theory calculations. These results pave the way toward the optimization of nanotube-based sensors with tailored sensitivity and selectivity to different species at room temperature. PMID:24404865

  12. 1H relaxation dispersion in solutions of nitroxide radicals: influence of electron spin relaxation.

    PubMed

    Kruk, D; Korpała, A; Kubica, A; Kowalewski, J; Rössler, E A; Moscicki, J

    2013-03-28

    The work presents a theory of nuclear ((1)H) spin-lattice relaxation dispersion for solutions of (15)N and (14)N radicals, including electron spin relaxation effects. The theory is a generalization of the approach presented by Kruk et al. [J. Chem. Phys. 137, 044512 (2012)]. The electron spin relaxation is attributed to the anisotropic part of the electron spin-nitrogen spin hyperfine interaction modulated by rotational dynamics of the paramagnetic molecule, and described by means of Redfield relaxation theory. The (1)H relaxation is caused by electron spin-proton spin dipole-dipole interactions which are modulated by relative translational motion of the solvent and solute molecules. The spectral density characterizing the translational dynamics is described by the force-free-hard-sphere model. The electronic relaxation influences the (1)H relaxation by contributing to the fluctuations of the inter-molecular dipolar interactions. The developed theory is tested against (1)H spin-lattice relaxation dispersion data for glycerol solutions of 4-oxo-TEMPO-d16-(15)N and 4-oxo-TEMPO-d16-(14)N covering the frequency range of 10 kHz-20 MHz. The studies are carried out as a function of temperature starting at 328 K and going down to 290 K. The theory gives a consistent overall interpretation of the experimental data for both (14)N and (15)N systems and explains the features of (1)H relaxation dispersion resulting from the electron spin relaxation.

  13. 1H relaxation dispersion in solutions of nitroxide radicals: Influence of electron spin relaxation

    NASA Astrophysics Data System (ADS)

    Kruk, D.; Korpała, A.; Kubica, A.; Kowalewski, J.; Rössler, E. A.; Moscicki, J.

    2013-03-01

    The work presents a theory of nuclear (1H) spin-lattice relaxation dispersion for solutions of 15N and 14N radicals, including electron spin relaxation effects. The theory is a generalization of the approach presented by Kruk et al. [J. Chem. Phys. 137, 044512 (2012)], 10.1063/1.4736854. The electron spin relaxation is attributed to the anisotropic part of the electron spin-nitrogen spin hyperfine interaction modulated by rotational dynamics of the paramagnetic molecule, and described by means of Redfield relaxation theory. The 1H relaxation is caused by electron spin-proton spin dipole-dipole interactions which are modulated by relative translational motion of the solvent and solute molecules. The spectral density characterizing the translational dynamics is described by the force-free-hard-sphere model. The electronic relaxation influences the 1H relaxation by contributing to the fluctuations of the inter-molecular dipolar interactions. The developed theory is tested against 1H spin-lattice relaxation dispersion data for glycerol solutions of 4-oxo-TEMPO-d16-15N and 4-oxo-TEMPO-d16-14N covering the frequency range of 10 kHz-20 MHz. The studies are carried out as a function of temperature starting at 328 K and going down to 290 K. The theory gives a consistent overall interpretation of the experimental data for both 14N and 15N systems and explains the features of 1H relaxation dispersion resulting from the electron spin relaxation.

  14. Synthesis and electrical conductivity studies of metal chloro and nitroxide group containing styrene butadiene rubber

    NASA Astrophysics Data System (ADS)

    Anilkumar, T.; Ramesan, M. T.

    2014-10-01

    The introduction of different functional group in SBR was done by a simple reaction between sodium nitrite and mercuric chloride in the presence of phase transfer catalyst. The attachment of chlorine and NO2 functional group in the double bond of the butadiene was monitored by FTIR and UV spectroscopy. The structure and morphology of chloro nitro SBR was studied by SEM and XRD. Flame retardency studies revealed that the chemical modification imparts better flame resistance to chemically modified SBR. AC conductivity and dielectric properties of chloro- nitro SBR was higher than that of SBR and conductivity increases with the level of chemical modification.

  15. Oral health and systemic inflammatory, cardiac and nitroxid biomarkers in hemodialysis patients.

    PubMed

    Cotič, J; Ferran, M; Karišik, J; Jerin, A; Pussinen, P-J; Nemec, A; Pavlica, Z; Buturović-Ponikvar, J; Petelin, M

    2017-07-01

    Periodontal diseases have systemic inflammatory effects and have been adversely associated with cardiovascular diseases, which are also the most frequent cause of death in the end-stage renal disease. The aim of this cross-sectional study was to investigate the oral health and serum biomarkers among the hemodialysis (HD) patients in Slovenia. 111 HD patients were periodontally examined and their sera were assayed for C reactive protein (CRP), cardiac troponin T (TnT), nitrite/nitrate (NOx) and antibody levels to A. actinomycetemcomitans and P. gingivalis. The association of oral health with systemic response was analyzed with Kruskal-Wallis test, Fisher's exact test and multivariate linear regression. Bleeding on probing without periodontal pockets was present in 5.2%, calculus without periodontal pockets in 42.1%, shallow periodontal pockets in 39.5% and deep periodontal pockets in 13.2% of dentate patients. There were 28.8% edentulous participants. 63.1% of the patients had CRP levels higher than 3 mg/L and 34.2% higher than 10 mg/L. TnT was detectable in all participants, with 25.2% exhibiting levels higher than 100 ng/L. The median level of NOx was 43.1 µmol/L. Participants with higher CRP were more likely to be edentulous and have higher TnT levels. A direct association of oral health with TnT or NOx was not detected. HD patients in Slovenia have compromised oral health and increased serum inflammatory and cardiac biomarkers. Edentulousness was an independent predictor for the increased CRP, indicating a need for improved dental care to retain the teeth as long as possible.

  16. Oral health and systemic inflammatory, cardiac and nitroxid biomarkers in hemodialysis patients

    PubMed Central

    Cotič, Jasna; Ferran, Monika; Karišik, Jasmina; Jerin, Aleš; Pussinen, Pirkko J.; Nemec, Ana; Pavlica, Zlatko; Buturović-Ponikvar, Jadranka

    2017-01-01

    Background Periodontal diseases have systemic inflammatory effects and have been adversely associated with cardiovascular diseases, which are also the most frequent cause of death in the end-stage renal disease. The aim of this cross-sectional study was to investigate the oral health and serum biomarkers among the hemodialysis (HD) patients in Slovenia. Material and Methods 111 HD patients were periodontally examined and their sera were assayed for C reactive protein (CRP), cardiac troponin T (TnT), nitrite/nitrate (NOx) and antibody levels to A. actinomycetemcomitans and P. gingivalis. The association of oral health with systemic response was analyzed with Kruskal-Wallis test, Fisher’s exact test and multivariate linear regression. Results Bleeding on probing without periodontal pockets was present in 5.2%, calculus without periodontal pockets in 42.1%, shallow periodontal pockets in 39.5% and deep periodontal pockets in 13.2% of dentate patients. There were 28.8% edentulous participants. 63.1% of the patients had CRP levels higher than 3 mg/L and 34.2% higher than 10 mg/L. TnT was detectable in all participants, with 25.2% exhibiting levels higher than 100 ng/L. The median level of NOx was 43.1 µmol/L. Participants with higher CRP were more likely to be edentulous and have higher TnT levels. A direct association of oral health with TnT or NOx was not detected. Conclusions HD patients in Slovenia have compromised oral health and increased serum inflammatory and cardiac biomarkers. Edentulousness was an independent predictor for the increased CRP, indicating a need for improved dental care to retain the teeth as long as possible. Key words:Periodontal diseases, edentulousness, C reactive protein, cardiac troponin T, nitric oxide. PMID:28578371

  17. Cysteine-Specific Labeling of Proteins with a Nitroxide Biradical for Dynamic Nuclear Polarization NMR.

    PubMed

    Voinov, Maxim A; Good, Daryl B; Ward, Meaghan E; Milikisiyants, Sergey; Marek, Antonin; Caporini, Marc A; Rosay, Melanie; Munro, Rachel A; Ljumovic, Milena; Brown, Leonid S; Ladizhansky, Vladimir; Smirnov, Alex I

    2015-08-13

    Dynamic nuclear polarization (DNP) enhances the signal in solid-state NMR of proteins by transferring polarization from electronic spins to the nuclear spins of interest. Typically, both the protein and an exogenous source of electronic spins, such as a biradical, are either codissolved or suspended and then frozen in a glycerol/water glassy matrix to achieve a homogeneous distribution. While the use of such a matrix protects the protein upon freezing, it also reduces the available sample volume (by ca. a factor of 4 in our experiments) and causes proportional NMR signal loss. Here we demonstrate an alternative approach that does not rely on dispersing the DNP agent in a glassy matrix. We synthesize a new biradical, ToSMTSL, which is based on the known DNP agent TOTAPOL, but also contains a thiol-specific methanethiosulfonate group to allow for incorporating this biradical into a protein in a site-directed manner. ToSMTSL was characterized by EPR and tested for DNP of a heptahelical transmembrane protein, Anabaena sensory rhodopsin (ASR), by covalent modification of solvent-exposed cysteine residues in two (15)N-labeled ASR mutants. DNP enhancements were measured at 400 MHz/263 GHz NMR/EPR frequencies for a series of samples prepared in deuterated and protonated buffers and with varied biradical/protein ratios. While the maximum DNP enhancement of 15 obtained in these samples is comparable to that observed for an ASR sample cosuspended with ~17 mM TOTAPOL in a glycerol-d8/D2O/H2O matrix, the achievable sensitivity would be 4-fold greater due to the gain in the filling factor. We anticipate that the DNP enhancements could be further improved by optimizing the biradical structure. The use of covalently attached biradicals would broaden the applicability of DNP NMR to structural studies of proteins.

  18. Saturation Recovery EPR and Nitroxide Spin Labeling for Exploring Structure and Dynamics in Proteins.

    PubMed

    Yang, Zhongyu; Bridges, Michael; Lerch, Michael T; Altenbach, Christian; Hubbell, Wayne L

    2015-01-01

    Experimental techniques capable of determining the structure and dynamics of proteins are continuously being developed in order to understand protein function. Among existing methods, site-directed spin labeling in combination with saturation recovery (SR) electron paramagnetic resonance spectroscopy contributes uniquely to the determination of secondary and tertiary protein structure under physiological conditions, independent of molecular weight and complexity. In addition, SR of spin labeled proteins was recently demonstrated to be sensitive to conformational exchange events with characteristic lifetimes on the order of μs, a time domain that presents a significant challenge to other spectroscopic techniques. In this chapter, we present the theoretical background necessary to understand the capabilities of SR as applied to spin labeled proteins, the instrumental requirements, and practical experimental considerations necessary to obtain interpretable data, and the use of SR to obtain information on protein: (1) secondary structure via solvent accessibility measurements, (2) tertiary structure using interspin distance measurements, and (3) conformational exchange. © 2015 Elsevier Inc. All rights reserved.

  19. Magnetic exchange interaction in gadolinium(III) complex having aliphatic nitroxide radical TEMPO

    NASA Astrophysics Data System (ADS)

    Nakamura, Takeshi; Ishida, Takayuki

    2016-02-01

    We synthesized a new compound, [Gd(hfac)3(MeOH)(TEMPO)] (TEMPO = 2,2,6,6-tetramethylpiperidin-1-oxyl; Hhfac = 1,1,1,5,5,5-hexafluoropentane-2,4-dione) with the metal/radical ratio of 1/1. This compound has an advantage in the magnetic analysis, because the exchange coupling system is described with a unique coupling parameter J, when compared to the structure and magnetic properties of the previous TEMPO and related complexes. The X-ray crystal structure analysis of [Gd(hfac)3(MeOH)(TEMPO)] revealed the N-O and Gd-O(N) bond lengths are 1.299(9) and 2.307(5) Å, respectively, and the Gd-O-N angle is 149.4(5)°. The magnetic study clarified the Gd3+-radical antiferromagnetic interaction of 2J/kB = -3.5(1) K.

  20. Fast redox of composite electrode of nitroxide radical polymer and carbon with polyacrylate binder

    NASA Astrophysics Data System (ADS)

    Komaba, Shinichi; Tanaka, Tatsuya; Ozeki, Tomoaki; Taki, Takayuki; Watanabe, Hiroaki; Tachikawa, Hiroyuki

    For organic radical batteries, poly(2,2,6,6-tetramethylpiperidinyloxy-4-yl methacrylate) (PTMA) has been reported as a promising positive electrode material. The PTMA/C composite electrode prepared with polyacrylate binder demonstrated the fast redox performance for the application to aprotic secondary batteries. When the variation in discharge capacities of the PTMA/C composite electrode was tested galvanostatically at 20 C rates, the electrode retained 96% of the initial capacity after 1000 cycles. This is attributed to the fact that the redox of PTMA is a simple reaction to form the oxoammonium salt doped with ClO 4 - anions in the electrolyte. When the PTMA/C composite electrode was discharged at different C rates, the electrode retained 81% of the theoretical capacity even at 50 C rates. This remarkably high rate capability originates from the fast electron-transfer kinetic of the 2,2,6,6-tetramethylpiperidine- N-oxyl (so-called TEMPO) radical, partially jelled polyacrylate binder, and the improved conductivity throughout the electrode by thoroughly mixing with carbon.