OC 68.5 - Neutrophil Extracellular Traps as a Mediator of Cerebral Ischemia and Reperfusion Injury in Ischemic Stroke

Background: Thromboinflammatory processes are involved in cerebral ischemia/reperfusion injury in ischemic stroke, likely with neutrophil extracellular traps (NETs) as a mediator. However, the role of NETs in cerebral ischemia/reperfusion injury is yet to be clarified. In addition, the link between von Willebrand factor (VWF) and neutrophil recruitment in the ischemic brain might play a role in thromboinflammation, possibly by the formation of NETs.

Aims: Therefore, our aim was to investigate the involvement of NETs in cerebral ischemia/reperfusion injury and to explore the potential regulation of NETosis by VWF and the VWF-cleaving protease ADAMTS13 in cerebral ischemia/reperfusion injury.

Methods: The filament-induced transient middle cerebral artery occlusion model was used to induce 60 minutes focal cerebral ischemia in wild-type (WT) and littermate Vwf-knockout (KO) and Adamts13-KO mice after which reperfusion was allowed. At different timepoints post-ischemia, NETs (Ly6G, H3Cit, DNA) were identified in the mouse brains using quantitative immunofluorescence microscopy.

Results: NETs could be identified in the ipsilateral brain hemisphere. Interestingly, quantification of NETs at different timepoints, revealed that NETs can already be detected at 6 hours, peak around 24 hours and started to disappear again 48 hours post-ischemia. Remarkably, NETs were predominantly localized within the brain vasculature at different timepoints (12 hours, 24 hours, 48 hours) post-ischemia and to a lesser extent in the brain parenchyma, suggesting that NETs might play a role in secondary microthrombosis. Strikingly, NET formation was significantly decreased in Vwf-KO mice and significantly increased in Adamts13-KO mice compared to littermate WT mice 24 hours post-ischemia.

Conclusion(s): Our results put forward NETs as a thromboinflammatory mediator and indicate a role for VWF in promoting NETosis in the ischemic brain. Taken together, better understanding of thromboinflammatory processes leading to cerebral ischemia/reperfusion injury in ischemic stroke could serve as a basis for the development of novel treatment strategies.