Abstract TP110: Polymerase δ-Interacting Protein 2 Regulates Astrocyte Activation in Ischemic Stroke

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Abstract

Introduction: Polymerase δ-Interacting Protein 2 (Poldip2) is a binding partner of Nox4 NADPH oxidase and carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1). Nox4 and CEACAM1 have been implicated in stroke, but with different outcomes: both contribute to neurodegeneration but only CEACAM1 contributes to blood brain barrier (BBB) dysfunction. However, the underlying mechanisms are still obscure.

Hypothesis: Poldip2+/- mice may be protected from the consequences of transient middle cerebral artery occlusion (tMCAO).

Methods: tMCAO was induced in wild type (WT) and Poldip2+/- mice. The volume of the ischemic lesion was measured in TTC-stained sections. BBB breakdown was evaluated by Evans blue dye extravasation. Poldip2 protein expression was evaluated by immunofluorescence and western blotting. RT-PCR was used to measure mRNA levels of cytokines, MMPs and TIMPs. Astrocytes were transfected with Poldip2 siRNA in culture and mRNA levels of cytokines were evaluated.

Results: Poldip2+/- and WT mice displayed comparable infarct sizes following tMCAO (n=6). A decrease in Evans blue dye extravasation was observed in Poldip2+/- mice (25±3 vs 6±2uM/g) (n=7). Upregulation of cytokine mRNA following tMCAO was also attenuated in Poldip2+/- mice (n=5): MCP-1(253±34 vs 83±23AU), IL-6 (134±38 vs 38±10AU), TNFα (39±12 vs 12±3AU), MMP-2 (19±3 vs 10±1AU), MMP-9 (253±12 vs 13±3AU) and TIMP-1 (286±110 vs 61±15AU). Poldip2 protein expression increased in the ischemic brain of WT mice after tMCAO (69±4 vs 20±5AU) and was predominantly located in astrocytes (n=4). Poldip2 protein expression was also increased in astrocytes following oxygen and glucose deprivation (79±15 vs 27±5AU) (n=6), and Poldip2 siRNA prevented cytokine induction under these conditions.

Conclusions: In conclusion, Poldip2 contributes to stroke-induced BBB breakdown via its ability to inhibit a pro- inflammatory response in perivascular astrocytes.

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