Abstract SNACC-69

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Peroxiredoxin-1 and Toll-like receptor 2 pathway contributes to neurotoxic microglial activation after cardiac arrest

Ikeda M, Fujiyoshi T, Mader S, Koerner I
Oregon Health and Science University, Portland, Oregon, United states

Introduction
Survivors of cardiac arrest (CA) and cardiopulmonary resuscitation (CPR) suffer cognitive decline related to delayed neuronal death in the hippocampus. We postulated that microglia are activated to a neurotoxic phenotype after CA/CPR, and that this contributes to delayed neuronal death. We tested whether danger signal peroxiredoxin-1 (Prx1) causes neurotoxic microglial activation after CA/CPR, and whether this requires microglial expression of TLR2.
Methods
Wild-type (WT) and TLR2-KO mice were subjected to CA/CPR. Neuronal death and microglial activation were evaluated 3 days later. Hippocampal tissue was harvested and cerebrospinal fluid (CSF) was collected from the foramen magnum for quantification of Prx1 protein. Microglia were isolated for RT-PCR analysis of TLR2-expression. Neurotoxicity of microglia was assessed in vitro by measuring neuronal death in microglia-neuronal co-cultures after oxygen-glucose deprivation (OGD). Group differences were evaluated using ANOVA or Student’s t-test, as appropriate. All data are presented as mean ±SEM.
Results
Microglial expression of TLR2 increased after CA/CPR (Naïve: 33.4±5.3, CA/CPR: 1day: 95.1±22.0, 7days: 64.3±10.9, Naïve vs 1day: P<0.01). Similarly, Prx1 protein expression was upregulated in mouse hippocampus after CA/CPR (Naïve: 0.27±0.04, CA/CPR: 2hrs: 0.58±0.12, 1day: 0.63±0.07, 3days: 0.44±0.04, Naïve vs 2hrs and 1day, P<0.05). Prx1 was released and detectable in the CSF within the first day after CA/CPR (see Fig. 1), followed by microglial activation and subsequent delayed neuronal death. Microglial activation was attenuated in TLR2-KO mice 3 days after CA/CPR (see Fig. 2), while neuronal death was not different (dead cell density in hippocampal CA1 area (cells/mm2), WT: 296±126, TLR2-KO: 620±310, P=0.36). Neuronal death in microglia-neuronal co-cultures after OGD increased when WT microglia were pretreated with Prx1 (WT microglia: 25±5 %, WT microglia+Prx1: 34±3 %, P<0.05), on the other hand, there was no significant increase in neuronal death when TLR2-KO microglia were pretreated with Prx1 (TLR2-KO microglia: 32±4 %, TLR2-KO microglia+Prx1: 34±3% P=0.46).
Conclusions
Microglia are activated to a neurotoxic phenotype after CA/CPR, which is facilitated by danger signal Prx1. WT, but not TLR2-KO microglia stimulated with Prx1 exacerbated neuronal death in vitro. Microglial activation after CA/CPR and Prx1 stimulation requires TLR2, whereas neurotoxicity is abolished by TLR2-deletion in vitro only. Further studies are needed to understand the differential role of TLR2 in microglial activation and neurotoxicity.

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