Leukemia Inhibitory Factor Protects Neurons from Ischemic Damage via Upregulation of Superoxide Dismutase 3.

Stephanie M Davis,Christopher C Leonardo,Keith R Pennypacker,Hilary A Seifert,Lisa A Collier,Craig T Ajmo

doi:10.1007/s12035-015-9587-2

Abstract

Leukemia inhibitory factor (LIF) has been shown to protect oligodendrocytes from ischemia by upregulating endogenous antioxidants. The goal of this study was to determine whether LIF protects neurons during stroke by upregulating superoxide dismutase 3 (SOD3). Animals were administered phosphate-buffered saline (PBS) or 125 μg/kg LIF at 6, 24, and 48 h after middle cerebral artery occlusion or sham surgery. Neurons were isolated from rat pups on embryonic day 18 and used between 7 and 15 days in culture. Cells were treated with LIF and/or 10 μM Akt inhibitor IV with PBS and 0.1 % DMSO acting as vehicle controls. Neurons transfected with scrambled or SOD3 small interfering RNA (siRNA) were subjected to 24-h ischemia after PBS or LIF treatment. LIF significantly increased superoxide dismutase activity and SOD3 expression in ipsilateral brain tissue compared to PBS. Following 24-h ischemia, LIF reduced cell death and increased SOD3 messenger RNA (mRNA) in vitro compared to PBS. Adding Akt inhibitor IV with LIF counteracted the decrease in cell death. Partially silencing the expression of SOD3 using siRNA prior to LIF treatment counteracted the protective effect of LIF-alone PBS treatment. These results indicate that LIF protects neurons in vivo and in vitro via upregulation of SOD3.

Highlights

Reactive oxygen species (ROS) play an instrumental role in both the acute and delayed phases of neuronal death during focal cerebral ischemia (FCI)
Total superoxide dismutase (SOD) activity was measured in brain lysates from rats euthanized 24, 48, or 72 h after middle cerebral artery occlusion (MCAO) or sham surgery
Mean SOD activities were normalized to the mean activity in samples from 72 h sham-operated rats

Summary

Introduction

Reactive oxygen species (ROS) play an instrumental role in both the acute and delayed phases of neuronal death during focal cerebral ischemia (FCI). During the early phase of ischemia, energy failure leads to membrane depolarization [1], calcium influx, and activation of pro-oxidant enzymes such as neuronal nitric oxide (NO) synthase [2] and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase [3]. The activation of microglia exacerbates oxidative damage to neurons via inducible NO synthase activation [6]. Since the therapeutic window for tissue plasminogen activator, the only FDA-approved drug for ischemic stroke, ranges from 3 to 4.5 h after stroke, it cannot target this delayed phase of neuronal death [12]. The need for novel stroke therapeutics that will protect vulnerable neurons from oxidative stress remains high

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