Our laboratory has shown that systemic inhibition of mitochondrial complex I with rotenone reproduces in rats and monkeys many features of PD, including dopaminergic degeneration, Lewy body and neurite (alpha-synuclein) pathology, DJ-i modification and translocation to mitochondria, and impairment of the ubiquitin-proteasome system. Most recently, we have found that there is iron deposition in substantia nigra of rotenone treated rats and monkeys, which appears identical to that seen in PD brains. In this model, transferrin (Tf) becomes oxidized (at Cys26o) and accumulates in nigral dopaminergic neurons. In collaboration with Charleen Chu, director of the Neuropathology Core, Greenamyre has found a similar - previously undescribed - accumulation of transferrin in nigral neurons in human postmortem specimens from PD cases. Additionally, he has found that transferrin receptor 2 (TfR2): (i) has a mitochondrial targeting sequence;(ii) is localized, in part, in mitochondria;and (iii) protein is selectively expressed in nigral dopamine neurons in rats.
Aim i : We will determine the sequence of cell types that accumulate iron after rotenone using the in vivo model and organotypic midbrain slice cultures, and we vAW ultimately assess this in human postmortem specimens across Braak staging.
Aim 2 : We will characterize (i) the oxidation state and distributions of Tf and TfR2 in the rotenone brain, (ii) the functional consequences of Tf oxidation, and (iii) ultimately, the distributions and oxidation states of these proteins in human specimens.
Aim 3 : We will assess the functional role of TfR2 by overexpressing or silencing the TfR2 gene under basal conditions and with rotenone treatment. Outcomes to be measured include: aspects of iron homeostasis, oxidative stress, a-synuclein/cytochrome c interactions, activation of cell death and protective mechanisms and cell viability.
Aim 4 : Lastly, we will manipulate - via viral-mediated gene transfer - proteins that regulate or chelate iron. We will look at the effects of manipulating ferritin and frataxin on rotenone-induced toxicity. We believe this project will elucidate (i) the mechanisms by which iron accumulates in PD and (ii) its role in PD pathogenesis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Program Projects (P01)
Project #
5P01NS059806-05
Application #
8505549
Study Section
National Institute of Neurological Disorders and Stroke Initial Review Group (NSD)
Project Start
Project End
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
5
Fiscal Year
2013
Total Cost
$253,422
Indirect Cost
$83,339
Name
University of Pittsburgh
Department
Type
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
An, Chengrui; Shi, Yejie; Li, Peiying et al. (2014) Molecular dialogs between the ischemic brain and the peripheral immune system: dualistic roles in injury and repair. Prog Neurobiol 115:6-24
Tapias, Victor; Greenamyre, J Timothy (2014) A rapid and sensitive automated image-based approach for in vitro and in vivo characterization of cell morphology and quantification of cell number and neurite architecture. Curr Protoc Cytom 68:12.33.1-12.33.22
Mohammadyani, Dariush; Tyurin, Vladimir A; O'Brien, Matthew et al. (2014) Molecular speciation and dynamics of oxidized triacylglycerols in lipid droplets: Mass spectrometry and coarse-grained simulations. Free Radic Biol Med 76:53-60
Hu, Xiaoming; Liou, Anthony K F; Leak, Rehana K et al. (2014) Neurobiology of microglial action in CNS injuries: receptor-mediated signaling mechanisms and functional roles. Prog Neurobiol 119-120:60-84
Tapias, Victor; Cannon, Jason R; Greenamyre, J Timothy (2014) Pomegranate juice exacerbates oxidative stress and nigrostriatal degeneration in Parkinson's disease. Neurobiol Aging 35:1162-76
Sanders, Laurie H; McCoy, Jennifer; Hu, Xiaoping et al. (2014) Mitochondrial DNA damage: molecular marker of vulnerable nigral neurons in Parkinson's disease. Neurobiol Dis 70:214-23
Sanders, Laurie H; Laganiere, Josee; Cooper, Oliver et al. (2014) LRRK2 mutations cause mitochondrial DNA damage in iPSC-derived neural cells from Parkinson's disease patients: reversal by gene correction. Neurobiol Dis 62:381-6
Wang, Jiayin; Shi, Yejie; Zhang, Lili et al. (2014) Omega-3 polyunsaturated fatty acids enhance cerebral angiogenesis and provide long-term protection after stroke. Neurobiol Dis 68:91-103
Tapias, Victor; Greenamyre, J Timothy; Watkins, Simon C (2013) Automated imaging system for fast quantitation of neurons, cell morphology and neurite morphometry in vivo and in vitro. Neurobiol Dis 54:158-68
Sanders, Laurie H; Greenamyre, J Timothy (2013) Oxidative damage to macromolecules in human Parkinson disease and the rotenone model. Free Radic Biol Med 62:111-20

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