RESEARCH ARTICLE Stat3 Mediates LIF-Induced Protection of Astrocytes Against Toxic ROS by Upregulating the UPC2 mRNA Pool Daniel W. Lapp, 1 Samuel S. Zhang, 1,2 and Colin J. Barnstable 1,2 Reactive oxygen species (ROS) have been implicated in various types of CNS damage, including stroke. We used a cultured astro- cyte model to explore mechanisms of survival of CNS cells following ROS damage. We found that pretreatment with leukemia inhibitory factor (LIF) preserves astrocytes exposed to toxic levels of t-BHP by inhibiting an increase in intracellular ROS following t- BHP treatment. Astrocytes lacking functional Stat3 did not benefit from the pro-survival or antioxidant effects of LIF. Inhibition of mitochondrial uncoupling protein 2 (UCP2) using a chemical inhibitor or siRNA abrogates the prosurvival effects of LIF, indicating a critical role for UCP2 in modulation of mitochondrial ROS production in survival following ROS exposure. LIF treatment of astro- cytes results in increased UCP2 mRNA that is accompanied by an increase in Stat3 binding to the UCP2 promoter region. Although treatment with LIF alone did not increase UCP2 protein, a combination of LIF treatment and ROS stress led to increased UCP2 pro- tein levels. We conclude that LIF protects astrocytes from ROS-induced death by increasing UCP2 mRNA, allowing cells to respond to ROS stress by rapidly producing UCP2 protein that ultimately decreases endogenous mitochondrial ROS production. GLIA 2014;62:159–170 Key words: glia, oxidative stress, uncoupling protein 2 Introduction I schemic damage in the central nervous system is a problem of critical importance due to the largely irreversible nature and the severe consequences of such injuries. Oxidative stress following ischemic injury is thought to be a major cause of subsequent pathology (Panickar and Norenberg, 2005; Siesj€ o et al., 1989). Glia play a critical role in the protection of neu- rons from reactive oxygen species (ROS)-mediated damage during central nervous system (CNS) insult (Barretto et al., 2011; Dienel and Hertz, 2005; Sofroniew, 2000). Reactive gliosis is a response of astrocytes to CNS injury, and involves changes in protein synthesis and morphology of astrocytes, as well as cytokine secretion by these cells (Banner et al., 1997; Ridet et al., 1997). While this process is often seen as detri- mental to CNS cell survival due to its ubiquitous presence in CNS injury, multiple lines of evidence demonstrate the pro- survival functions of astrocytes in the CNS (Desagher et al., 1996; Lucius and Severs, 1996; Macauley, 2011; Spence et al., 2011; Tanaka et al., 1999). The ability of glia to pre- vent ROS-induced death in the CNS is due in part to the production of antioxidant enzymes (Barretto et al., 2011). Astrocytes, therefore, contribute to survival of CNS tissues in the face of toxic ROS by both preventing ROS damage and by maintaining their normal functions. However, the mecha- nisms by which glia themselves survive damage from oxygen free radicals remain incompletely understood. Cells are susceptible to ROS damage when the overall oxidant burden exceeds the capacity of intracellular coping mechanisms to manage the negative effects. A cell’s oxidant bur- den is the sum of endogenously produced ROS, as well as ROS from exogenous sources or abnormal processes. To improve survival against oxidative stress a cell can either increase antioxidant defenses, or decrease endogenous produc- tion to lower total ROS burden to manageable levels. Mito- chondrial uncoupling proteins are known to disconnect the electron transport chain from oxidative phosphorylation (Hor- vath et al., 2003) resulting in a decrease of endogenous ROS production by the mitochondria (Negre-Salverye et al., 1997), View this article online at wileyonlinelibrary.com. DOI: 10.1002/glia.22594 Published online December 5, 2013 in Wiley Online Library (wileyonlinelibrary.com). Received Aug 1, 2013, Accepted for publication Oct 16, 2013. Address correspondence to Colin J. Barnstable, Department of Neural and Behavioral Sciences, Penn State Milton S. Hershey Medical Center, 500 University Drive, Hershey, PA 17033 USA. E-mail: cjb30@psu.edu From the 1 Department of Neural and Behavioral Sciences, Penn State College of Medicine, Hershey, Pennsylvania; 2 Penn State Hershey Eye Center, Penn State College of Medicine, Hershey, Pennsylvania. V C 2013 Wiley Periodicals, Inc. 159