Injury to the brain caused by cerebral ischemia is a major public health concern. Studies have determined that the brain damage associated with cerebral ischemia is mediated by over-stimulation of excitatory amino acid receptors, oxidative stress, as well as inflammatory factors. During the last grant period our laboratory demonstrated that astrocyte-mediated alterations in system xc- (cystine/glutamate antiporter) activity contributes to the development and progression of inflammatory (IL-1beta-enhanced) hypoxic neuronal injury -a model of the ischemic penumbra. Despite this, new preliminary data demonstrate that IL-1beta-mediated upregulation of the same molecule, system xc-, can confer protection against oxidative insults. We speculate that IL-1beta upregulation of astrocyte system xc- may have evolved as a protective mechanism to counteract oxidative stress in injured tissue. However, this increase becomes maladaptive in the setting of compromised glutamate uptake, which occurs in the setting of our hypoxia model in vitro and stroke in vivo. The concept that IL-1beta and system xc- are at the crossroads of injury and protection is particularly intriguing. Studies to systematically and empirically address these ideas, as well as, to elucidate the regulation of the transporter by IL-1beta at the molecular level are solely needed. Thus, the objectives of this following 5 yr research plan of study are as follows: 1) To determine the mechanism by which IL-1beta regulates astrocyte system xc- expression. State of the art molecular biological approaches will be utilized to assess whether IL-1beta regulates xCT mRNA at the transcriptional and/or post-transcriptional level and to identify the cis and trans-acting factors responsible for the induction and/or stabilization of xCT message. 2) To determine the functional consequence of enhanced system xc- activity. The goal of this aim is to determine whether the IL-1beta-mediated enhancement of system xc- activity, a priori, increases GSH content and confers a selective resistance to oxidative injury under conditions where glutamate uptake is competent both in vitro and in vivo. 3) Using genetic approaches, studies will be undertaken to determine the extent to which IL-1beta signaling regulates system xc- expression following cerebral ischemic injury with the direct question as to whether loss of system xc- function either globally, or in astrocytes specifically, can alter the susceptibility of mouse brai to cerebral ischemic damage. Understanding the regulation of system xc- by IL-1beta is necessary so that we may use this information to devise strategies to harness the beneficial effects (i.e. to increase GSH levels to reduce oxidative injury), and when appropriate, to employ strategies to reduce its activity to decrease the probability of excitotoxic neuronal injury (i.e. under conditios where glutamate uptake is impaired).
Morbidity associated with stroke remains a huge emotional and economic burden due in large part to a void in treatment options to protect against secondary injury. It is our contention that successful completion of this proposal will advance and refine our knowledge about an important new therapeutic target (system xc-) that complements other ongoing efforts to reduce injury following cerebral ischemic insult.
|Lewerenz, Jan; Hewett, Sandra J; Huang, Ying et al. (2013) The cystine/glutamate antiporter system x(c)(-) in health and disease: from molecular mechanisms to novel therapeutic opportunities. Antioxid Redox Signal 18:522-55|
|Hewett, James A; Hewett, Sandra J (2012) Induction of nitric oxide synthase-2 expression and measurement of nitric oxide production in enriched primary cortical astrocyte cultures. Methods Mol Biol 814:251-63|
|Claycomb, Robert J; Hewett, Sandra J; Hewett, James A (2012) Neuromodulatory role of endogenous interleukin-1ýý in acute seizures: possible contribution of cyclooxygenase-2. Neurobiol Dis 45:234-42|
|Jackman, Nicole A; Melchior, Shannon E; Hewett, James A et al. (2012) Non-cell autonomous influence of the astrocyte system xc- on hypoglycaemic neuronal cell death. ASN Neuro 4:|
|Uliasz, Tracy F; Hamby, Mary E; Jackman, Nicole A et al. (2012) Generation of primary astrocyte cultures devoid of contaminating microglia. Methods Mol Biol 814:61-79|
|Jackman, Nicole A; Uliasz, Tracy F; Hewett, James A et al. (2010) Regulation of system x(c)(-)activity and expression in astrocytes by interleukin-1?: implications for hypoxic neuronal injury. Glia 58:1806-15|
|Hamby, Mary E; Hewett, James A; Hewett, Sandra J (2010) Smad3-dependent signaling underlies the TGF-ýý1-mediated enhancement in astrocytic iNOS expression. Glia 58:1282-91|
|Fogal, Birgit; Hewett, Sandra J (2008) Interleukin-1beta: a bridge between inflammation and excitotoxicity? J Neurochem 106:1-23|
|Fogal, Birgit; Li, Jun; Lobner, Doug et al. (2007) System x(c)- activity and astrocytes are necessary for interleukin-1 beta-mediated hypoxic neuronal injury. J Neurosci 27:10094-105|