Tissue plasminogen activator (tPA) thrombolysis is beneficial for thrombotic stroke, but its direct neuronal and vascular toxicities are problematic. Our extensive pilot data suggest that: (1) protein S (PS) has antithrombotic and neuroprotective activities in mouse stroke models; (2) PS protects neurons and brain endothelial cells (BEC) from N-methyl-D-aspartate (NMDA) and oxygen/glucose deprivation (OGD) injuries; (3) PS's cytoprotection requires its C-terminal sex hormone binding globulin-like (rSHBG) module, but not the N-terminal micro-PS [Gla domain, thrombin-sensitive (TSR) region, EGF1 module]; (4) PS activates the PISK-Akt cell survival axis and inhibits the intrinsic apoptotic cascade in NMDA-treated neurons and OGD- treated BEC and the extrinsic apoptotic cascade in tPA/NMDA-treated neurons and tPA/OGD-treated BEC; and (5) PS's cytoprotection may require the rse/Tyro-3 receptor tyrosine kinase (RTK), but not the Axl/Tyro 7 or Mer/Tyro 12 RTKs. Thus, we hypothesize that PS and certain of its variants may protect brain from ischemic/thrombotic events by antithrombotic activity involving Gla, TSR and EGF1 modules, and by direct cytoprotection involving the SHBG domain. We further hypothesize that combined therapies for stroke with tPA and PS, rSHBG and the GlaFII PS mutant will directly protect brain cells because PS and its variants with intact SHBG domain can activate the Tyro 3-Akt cell survival signaling pathway, and because this cytoprotective action can compensate for tPA's cytotoxicities. The research design proposes to test the hypothesis: first, using mechanical and embolic mouse stroke models (aim 1); second, using an in vivo mouse model of NMDA and tPA excitotoxic brain lesions to isolate intravascular PS and tPA effects from their direct effects on brain cells (aim 2); and third, using in vitro models of NMDA and tPA/NMDA neuronal injuries (aim 3) and OGD and tPA/OGD BEC injuries (aim 4). Expertise and reagents provided by collaborators/consultants will be Dr. J.H. Griffin (Scripps) (PS reagents, mouse tPA), Dr. G. Lemke (Salk) (Tyro 3 RTK mutants), Dr. R. Freeman (Rochester) (various Akt, Bel and FKHRL1 mutants), Dr. M. Moskowitz (Harvard) (intrinsic/extrinsic apoptotic signaling), Dr. B. Berk (Rochester) (Tyro 3 RTK-Akt signaling) and Dr. M. Chopp (Henry Ford) (rat stroke model). The results will provide mechanistic insights and test therapeutic agents that may be translated to improved therapy for ischemic stroke. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL081528-03
Application #
7457899
Study Section
Clinical Neuroscience and Disease Study Section (CND)
Program Officer
Kindzelski, Andrei L
Project Start
2006-07-01
Project End
2011-04-30
Budget Start
2008-05-01
Budget End
2009-04-30
Support Year
3
Fiscal Year
2008
Total Cost
$378,690
Indirect Cost
Name
University of Rochester
Department
Neurosurgery
Type
Schools of Dentistry
DUNS #
041294109
City
Rochester
State
NY
Country
United States
Zip Code
14627
Zlokovic, Berislav V; Griffin, John H (2011) Cytoprotective protein C pathways and implications for stroke and neurological disorders. Trends Neurosci 34:198-209
Zhu, Donghui; Wang, Yaoming; Singh, Itender et al. (2010) Protein S controls hypoxic/ischemic blood-brain barrier disruption through the TAM receptor Tyro3 and sphingosine 1-phosphate receptor. Blood 115:4963-72
Walker, Corey T; Marky, Andrew H; Petraglia, Anthony L et al. (2010) Activated protein C analog with reduced anticoagulant activity improves functional recovery and reduces bleeding risk following controlled cortical impact. Brain Res 1347:125-31
Petraglia, Anthony L; Marky, Andrew H; Walker, Corey et al. (2010) Activated protein C is neuroprotective and mediates new blood vessel formation and neurogenesis after controlled cortical impact. Neurosurgery 66:165-71; discussion 171-2
Zhong, Zhihui; Wang, Yaoming; Guo, Huang et al. (2010) Protein S protects neurons from excitotoxic injury by activating the TAM receptor Tyro3-phosphatidylinositol 3-kinase-Akt pathway through its sex hormone-binding globulin-like region. J Neurosci 30:15521-34
Deane, Rashid; LaRue, Barbra; Sagare, Abhay P et al. (2009) Endothelial protein C receptor-assisted transport of activated protein C across the mouse blood-brain barrier. J Cereb Blood Flow Metab 29:25-33
Zhong, Zhihui; Ilieva, Hristelina; Hallagan, Lee et al. (2009) Activated protein C therapy slows ALS-like disease in mice by transcriptionally inhibiting SOD1 in motor neurons and microglia cells. J Clin Invest 119:3437-49
Fernandez, Jose A; Heeb, Mary J; Xu, Xiao et al. (2009) Species-specific anticoagulant and mitogenic activities of murine protein S. Haematologica 94:1721-31
Guo, Huang; Singh, Itender; Wang, Yaoming et al. (2009) Neuroprotective activities of activated protein C mutant with reduced anticoagulant activity. Eur J Neurosci 29:1119-30
Guo, Huang; Wang, Yaoming; Singh, Itender et al. (2009) Species-dependent neuroprotection by activated protein C mutants with reduced anticoagulant activity. J Neurochem 109:116-24

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