An important mechanism of ischemic brain damage is production of reactive O2 species, including superoxide (O2-). Natural defenses against O2- include 3 isozymes of superoxide dismutase (SOD). CuZnSOD and MnSOD have been shown to be important in scavenging O2- produced in the intracellular space. The remaining isozyme, extracellular SOD (EC-SOD), is found only in the extracellular (EC) space. Transgenic (EC-SOD overexpressing) and knockout (EC-SOD deficient) mice and the metalloporphyrin compound Mn-TM-2-PyP (an EC-SOD mimetic) provide novel opportunity to isolate and examine effects of O2- produced in the EC space. We have shown that EC-SOD overexpression reduces both global and focal ischemic injury while EC-SOD deficiency increases focal ischemic injury. The goal of this research is to define mechanisms by which EC-SOD improves ischemic outcome and determine if these properties can be emulated by use of EC-SOD mimetic compounds. Our fundamental postulate is that EC-SOD provides a beneficial effect on ischemic brain by scavenging O2- in the EC space, generated by either activated neutrophils/microglia or membrane bound oxidases. We will examine whether reperfusion is required for EC-SOD to affect histologic/behavioral outcome. We will then examine whether EC-SOD deficiency worsens global ischemic injury and whether this deficiency can be corrected by administration of Mn-TM-2-PyP. O2- sensitive microelectrodes, OH-salicylate trapping by microdialysis, and nitrotyrosine assays will be used to determine if manipulation of EC-SOD expression and use of Mn-TM-2-PyP alters O2 concentrations in ischemic brain. The source of extracellular O2- will be examined by comparing histologic/behavioral ischemic outcome in neutrophil depleted EC-SOD transgenic/knockout mice and whether EC-SOD pharmacologic mimetics have efficacy in NADPH oxidase knock-out mice lacking respiratory burst activity. Long-term recovery studies will be performed to assure that neuroprotective effects of EC-SOD are permanent. Immunoblotting techniques will be used to determine if EC-SOD expression is upregulated in post-ischemic brain and whether expressed EC-SOD is intact or cleaved of its heparin-binding domain, which might facilitate diffusion to sites of inflammation. We believe this work will provide important mechanistic insight into how ischemic outcome is affected by O2- formed in the extracellular space and that this work will provide novel routes of investigation for therapy of ischemic brain injury.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Program Projects (P01)
Project #
2P01HL042444-11
Application #
6545648
Study Section
Heart, Lung, and Blood Program Project Review Committee (HLBP)
Project Start
1990-04-01
Project End
2006-06-30
Budget Start
Budget End
Support Year
11
Fiscal Year
2001
Total Cost
Indirect Cost
Name
Duke University
Department
Type
DUNS #
071723621
City
Durham
State
NC
Country
United States
Zip Code
27705
Schwab, David E; Stamler, Jonathan S; Singel, David J (2010) EPR spectroscopy of nitrite complexes of methemoglobin. Inorg Chem 49:6330-7
Sheng, Huaxin; Yang, Wei; Fukuda, Shiro et al. (2009) Long-term neuroprotection from a potent redox-modulating metalloporphyrin in the rat. Free Radic Biol Med 47:917-23
Zhu, Jun; Li, Sheng; Marshall, Zermeena M et al. (2008) A cystine-cysteine shuttle mediated by xCT facilitates cellular responses to S-nitrosoalbumin. Am J Physiol Cell Physiol 294:C1012-20
Buckley, Barbara J; Li, Sheng; Whorton, A Richard (2008) Keap1 modification and nuclear accumulation in response to S-nitrosocysteine. Free Radic Biol Med 44:692-8
Diesen, Diana L; Hess, Douglas T; Stamler, Jonathan S (2008) Hypoxic vasodilation by red blood cells: evidence for an s-nitrosothiol-based signal. Circ Res 103:545-53
Granillo, Olivia M; Brahmajothi, Mulugu V; Li, Sheng et al. (2008) Pulmonary alveolar epithelial uptake of S-nitrosothiols is regulated by L-type amino acid transporter. Am J Physiol Lung Cell Mol Physiol 295:L38-43
Gutsaeva, Diana R; Carraway, Martha Sue; Suliman, Hagir B et al. (2008) Transient hypoxia stimulates mitochondrial biogenesis in brain subcortex by a neuronal nitric oxide synthase-dependent mechanism. J Neurosci 28:2015-24
Reynolds, James D; Ahearn, Gregory S; Angelo, Michael et al. (2007) S-nitrosohemoglobin deficiency: a mechanism for loss of physiological activity in banked blood. Proc Natl Acad Sci U S A 104:17058-62
Nozik-Grayck, Eva; Whalen, Erin J; Stamler, Jonathan S et al. (2006) S-nitrosoglutathione inhibits alpha1-adrenergic receptor-mediated vasoconstriction and ligand binding in pulmonary artery. Am J Physiol Lung Cell Mol Physiol 290:L136-43
Leinenweber, Stephen B; Sheng, Huaxin; Lynch, John R et al. (2006) Effects of a manganese (III) porphyrin catalytic antioxidant in a mouse closed head injury model. Eur J Pharmacol 531:126-32

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