Persistent Link:
http://hdl.handle.net/10150/195215
Title:
Reactive Oxygen Species-Induced Necrotic Cell Death
Author:
Xie, Ruiyu
Issue Date:
2009
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Mechanisms of cell death extend beyond the simple apoptosis/necrosis relationship to include regulated modes of cell death that do not readily fit either of the classic descriptors. One such mechanism of cell death involves poly(ADP-ribose)polymerase-1 (PARP-1)-mediated cell death. 2,3,5-Tris(Glutathion-S-yl)-hydroquinone (TGHQ), a reactive oxygen species (ROS) generating nephrotoxic and nephrocarcinogenic metabolite of hydroquinone, causes necrotic renal cell death, the basis for which is unclear. We therefore investigated TGHQ-mediated cell death in human renal proximal tubule epithelial HK-2 cells. TGHQ induced ROS generation, DNA strand breaks, hyperactivation of PARP-1, rapid depletion of nicotinamide adenine dinucleotide (NAD), elevations in intracellular Ca2+ concentrations, loss of mitochondrial membrane potential, and subsequent necrotic cell death. Interestingly, PARP-1 hyperactivation was not accompanied by the translocation of apoptosis-inducing factor (AIF) from mitochondria to the nucleus, a process usually associated with PARP-dependent cell death. Inhibition of PARP-1 with PJ34 blocked TGHQ-mediated accumulation of poly(ADP-ribose) polymers, NAD consumption, and the consequent necrotic cell death. However, HK-2 cell death was only delayed by PJ34, and cell death remained necrotic in nature. In contrast, chelation of intracellular Ca2+ with BAPTA-AM completely abrogated TGHQ-induced necrotic cell death. Ca2+ chelation not only prevented the collapse in the mitochondrial potential but also attenuated PARP-1 hyperactivation. Conversely, inhibition of PARP-1 modulated TGHQ-mediated changes in Ca2+ homeostasis. Moreover, TGHQ caused a sequential oxidation of peroxiredoxin III (PrxIII), a protein considered the primary antioxidant defense within mitochondria. Thus, TGHQ induced two acidic shifts in PrxIII, with both pI shifted spots representing oxidized forms of PrxIII. Transient expression of a dominant negative version of PrxIII resulted in a significant increase in TGHQ-induced cytotoxicity, whereas overexpression of wild-type PrxIII significantly attenuated cytotoxicity. Our studies provide new insights into PARP-1-mediated necrotic cell death. Changes in intracellular Ca2+ concentrations appear to couple PARP-1-hyperactivation to subsequent cell death, but in the absence of AIF release from mitochondria. NAD depletion, mitochondrial membrane depolarization, Ca2+-mediated calpain activation, and PrxIII oxidation, all contribute to TGHQ-driven ROS-mediated necrotic cell death.
Type:
text; Electronic Dissertation
Keywords:
calcium; cell death; mechanism; necrosis; PARP; reactive oxygen species
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Pharmacology & Toxicology; Graduate College
Degree Grantor:
University of Arizona
Advisor:
Monks, Terrence J

Full metadata record

DC FieldValue Language
dc.language.isoENen_US
dc.titleReactive Oxygen Species-Induced Necrotic Cell Deathen_US
dc.creatorXie, Ruiyuen_US
dc.contributor.authorXie, Ruiyuen_US
dc.date.issued2009en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractMechanisms of cell death extend beyond the simple apoptosis/necrosis relationship to include regulated modes of cell death that do not readily fit either of the classic descriptors. One such mechanism of cell death involves poly(ADP-ribose)polymerase-1 (PARP-1)-mediated cell death. 2,3,5-Tris(Glutathion-S-yl)-hydroquinone (TGHQ), a reactive oxygen species (ROS) generating nephrotoxic and nephrocarcinogenic metabolite of hydroquinone, causes necrotic renal cell death, the basis for which is unclear. We therefore investigated TGHQ-mediated cell death in human renal proximal tubule epithelial HK-2 cells. TGHQ induced ROS generation, DNA strand breaks, hyperactivation of PARP-1, rapid depletion of nicotinamide adenine dinucleotide (NAD), elevations in intracellular Ca2+ concentrations, loss of mitochondrial membrane potential, and subsequent necrotic cell death. Interestingly, PARP-1 hyperactivation was not accompanied by the translocation of apoptosis-inducing factor (AIF) from mitochondria to the nucleus, a process usually associated with PARP-dependent cell death. Inhibition of PARP-1 with PJ34 blocked TGHQ-mediated accumulation of poly(ADP-ribose) polymers, NAD consumption, and the consequent necrotic cell death. However, HK-2 cell death was only delayed by PJ34, and cell death remained necrotic in nature. In contrast, chelation of intracellular Ca2+ with BAPTA-AM completely abrogated TGHQ-induced necrotic cell death. Ca2+ chelation not only prevented the collapse in the mitochondrial potential but also attenuated PARP-1 hyperactivation. Conversely, inhibition of PARP-1 modulated TGHQ-mediated changes in Ca2+ homeostasis. Moreover, TGHQ caused a sequential oxidation of peroxiredoxin III (PrxIII), a protein considered the primary antioxidant defense within mitochondria. Thus, TGHQ induced two acidic shifts in PrxIII, with both pI shifted spots representing oxidized forms of PrxIII. Transient expression of a dominant negative version of PrxIII resulted in a significant increase in TGHQ-induced cytotoxicity, whereas overexpression of wild-type PrxIII significantly attenuated cytotoxicity. Our studies provide new insights into PARP-1-mediated necrotic cell death. Changes in intracellular Ca2+ concentrations appear to couple PARP-1-hyperactivation to subsequent cell death, but in the absence of AIF release from mitochondria. NAD depletion, mitochondrial membrane depolarization, Ca2+-mediated calpain activation, and PrxIII oxidation, all contribute to TGHQ-driven ROS-mediated necrotic cell death.en_US
dc.typetexten_US
dc.typeElectronic Dissertationen_US
dc.subjectcalciumen_US
dc.subjectcell deathen_US
dc.subjectmechanismen_US
dc.subjectnecrosisen_US
dc.subjectPARPen_US
dc.subjectreactive oxygen speciesen_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplinePharmacology & Toxicologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorMonks, Terrence Jen_US
dc.contributor.committeememberJacobson, Myron Ken_US
dc.contributor.committeememberLau, Serrine Sen_US
dc.contributor.committeememberChen, Qin Men_US
dc.contributor.committeememberFutsher, Bernard Wen_US
dc.identifier.proquest10432en_US
dc.identifier.oclc659752064en_US
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