title: Post-traumatic administration of the p53 inactivator pifithrin-α oxygen analogue reduces hippocampal neuronal loss and improves cognitive deficits after experimental traumatic brain injury.
abstract: Traumatic brain injury (TBI) is a major cause of death and disability worldwide. Neuronal apoptosis in the hippocampus has been detected after TBI. The hippocampal dysfunction may result in cognitive deficits in learning, memory, and spatial information processing. Our previous studies demonstrated that a p53 inhibitor, pifithrin-α oxygen analogue (PFT-α (O)), significantly reduced cortical cell death, which is substantial following controlled cortical impact (CCI) TBI, and improved neurological functional outcomes via anti-apoptotic mechanisms. In the present study, we examined the effect of PFT-α (O) on CCI TBI-induced hippocampal cellular pathophysiology in light of this brain region's role in memory. To investigate whether p53-dependent apoptosis plays a role in hippocampal neuronal loss and associated cognitive deficits and to define underlying mechanisms, SD rats were subjected to experimental CCI TBI followed by the administration of PFT-α or PFT-α (O) (2mg/kg, i.v.) or vehicle at 5h after TBI. Magnetic resonance imaging (MRI) scans were acquired at 24h and 7days post-injury to assess evolving structural hippocampal damage. Fluoro-Jade C was used to stain hippocampal sub-regions, including CA1 and dentate gyrus (DG), for cellular degeneration. Neurological functions, including motor and recognition memory, were assessed by behavioral tests at 7days post injury. p53, p53 upregulated modulator of apoptosis (PUMA), 4-hydroxynonenal (4-HNE), cyclooxygenase-IV (COX IV), annexin V and NeuN were visualized by double immunofluorescence staining with cell-specific markers. Levels of mRNA encoding for caspase-3, p53, PUMA, Bcl-2, Bcl-2-associated X protein (BAX) and superoxide dismutase (SOD) were measured by RT-qPCR. Our results showed that post-injury administration of PFT-α and, particularly, PFT-α (O) at 5h dramatically reduced injury volumes in the ipsilateral hippocampus, improved motor outcomes, and ameliorated cognitive deficits at 7days after TBI, as evaluated by novel object recognition and open-field test. PFT-α and especially PFT-α (O) significantly reduced the number of FJC-positive cells in hippocampus CA1 and DG subregions, versus vehicle treatment, and significantly decreased caspase-3 and PUMA mRNA expression. PFT-α (O), but not PFT-α, treatment significantly lowered p53 and elevated SOD2 mRNA expression. Double immunofluorescence staining demonstrated that PFT-α (O) treatment decreased p53, annexin V and 4-HNE positive neurons in the hippocampal CA1 region. Furthermore, PUMA co-localization with the mitochondrial maker COX IV, and the upregulation of PUMA were inhibited by PFT-α (O) after TBI. Our data suggest that PFT-α and especially PFT-α (O) significantly reduce hippocampal neuronal degeneration, and ameliorate neurological and cognitive deficits in vivo via antiapoptotic and antioxidative properties.
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Ling-Yu
authors.initials: LY
authors.lastname: Yang
authors.affiliation: Drug Design & Development Section, Translational Gerontology Branch, Intramural Research Program, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
authors.forename: Nigel H
authors.initials: NH
authors.lastname: Greig
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan; Department of Nursing, Hsin Sheng Junior College of Medical Care and Management, Taoyuan, Taiwan.
authors.forename: Ya-Ni
authors.initials: YN
authors.lastname: Huang
authors.affiliation: Department of Physical Therapy and Graduate Institute of Rehabilitation Science, College of Medicine, Chang Gung University, Taoyuan, Taiwan; Graduate Institute of Neural Regenerative Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Tsung-Hsun
authors.initials: TH
authors.lastname: Hsieh
authors.affiliation: Drug Design & Development Section, Translational Gerontology Branch, Intramural Research Program, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
authors.forename: David
authors.initials: D
authors.lastname: Tweedie
authors.affiliation: Drug Design & Development Section, Translational Gerontology Branch, Intramural Research Program, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
authors.forename: Qian-Sheng
authors.initials: QS
authors.lastname: Yu
authors.affiliation: Department of Neurosurgery, Case Western Reserve University School of Medicine, Cleveland, OH, USA.
authors.forename: Barry J
authors.initials: BJ
authors.lastname: Hoffer
authors.affiliation: Department of Neurosurgery, Case Western Reserve University School of Medicine, Cleveland, OH, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Luo
authors.affiliation: Translational Imaging Research Center and Department of Radiology, School of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Yu-Chieh
authors.initials: YC
authors.lastname: Kao
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan; Department of Physiology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan. Electronic address: jywang2010@tmu.edu.tw.
authors.forename: Jia-Yi
authors.initials: JY
authors.lastname: Wang
mesh_terms: D000447:Aldehydes / Q000378:metabolism; D000818:Animals; D017304:Annexin A5 / Q000235:genetics / Q000378:metabolism; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D052160:Benzothiazoles / Q000737:chemistry / Q000494:pharmacology / Q000627:therapeutic use*; D001921:Brain / Q000000981:diagnostic imaging; D000070642:Brain Injuries, Traumatic / Q000150:complications* / Q000000981:diagnostic imaging; D003072:Cognition Disorders / Q000000981:diagnostic imaging / Q000188:drug therapy / Q000209:etiology / Q000473:pathology; D004195:Disease Models, Animal; D003576:Electron Transport Complex IV / Q000378:metabolism; D005106:Exploratory Behavior / Q000187:drug effects; D005452:Fluoresceins / Q000378:metabolism; D008279:Magnetic Resonance Imaging; D008297:Male; D010100:Oxygen; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D021641:Recognition, Psychology / Q000187:drug effects; D013997:Time Factors; D014050:Toluene / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology / Q000627:therapeutic use; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
chemical_list: D000447:Aldehydes; D017304:Annexin A5; D051017:Apoptosis Regulatory Proteins; C533952:Bbc3 protein, rat; D052160:Benzothiazoles; D005452:Fluoresceins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C435731:fluoro jade; D014050:Toluene; C121565:pifithrin; C548296:COX15 protein, human; D003576:Electron Transport Complex IV; C027576:4-hydroxy-2-nonenal; D010100:Oxygen
keywords: Apoptosis; Controlled cortical impact (CCI); Motor and cognitive deficits; PFT-α oxygen analogue (PFT-α (O)); Pifithrin-α (PFT-α); Puma; Traumatic brain injury (TBI); p53