Erythropoietin Abrogates Post-Ischemic Activation of the NLRP3, NLRC4, and AIM2 Inflammasomes in Microglia/Macrophages in a TAK1-Dependent Manner

Ole Heinisch,Eren Arik,Arno Reich,Shahin Habib,Marco Prinz,Jörg B Schulz,Pardes Habib,Michael Huber,Alexander Slowik,Tobias Goldmann,Thomas Zeyen,Jennifer Jung

doi:10.1007/s12975-021-00948-8

Abstract

Inflammasomes are known to contribute to brain damage after acute ischemic stroke (AIS). TAK1 is predominantly expressed in microglial cells and can regulate the NLRP3 inflammasome, but its impact on other inflammasomes including NLRC4 and AIM2 after AIS remains elusive. EPO has been shown to reduce NLRP3 protein levels in different disease models. Whether EPO-mediated neuroprotection after AIS is conveyed via an EPO/TAK1/inflammasome axis in microglia remains to be clarified. Subjecting mice deficient for TAK1 in microglia/macrophages (Mi/MΦ) to AIS revealed a significant reduction in infarct sizes and neurological impairments compared to the corresponding controls. Post-ischemic increased activation of TAK1, NLRP3, NLRC4, and AIM2 inflammasomes including their associated downstream cascades were markedly reduced upon deletion of Mi/MΦ TAK1. EPO administration improved clinical outcomes and dampened stroke-induced activation of TAK1 and inflammasome cascades, which was not evident after the deletion of Mi/MΦ TAK1. Pharmacological inhibition of NLRP3 in microglial BV-2 cells did not influence post-OGD IL-1β levels, but increased NLRC4 and AIM2 protein levels, suggesting compensatory activities among inflammasomes. Overall, we provide evidence that Mi/MΦ TAK1 regulates the expression and activation of the NLRP3, NLRC4, AIM2 inflammasomes. Furthermore, EPO mitigated stroke-induced activation of TAK1 and inflammasomes, indicating that EPO conveyed neuroprotection might be mediated via an EPO/TAK1/inflammasome axis.Graphical

Highlights

Acute ischemic stroke (AIS) remains a leading cause of death and long-term disability worldwide and is associated with a high socioeconomic burden [1,2,3]
Since little to nothing is known about the impact of EPO on microglia, microglial TGFβ-activated kinase 1 (TAK1), and on the inflammasomes after stroke, we deemed an investigation of the reperfusion time point of 6 h after 30 min of transient middle cerebral artery occlusion (tMCAo) essential to draw any conclusion about a potential EPO/TAK1/inflammasome axis
The subacute reperfusion phase of 72 h after stroke was chosen on the basis of our preliminary data, which showed a significant reduction in infarct volumes after this period upon three administrations of EPO

Summary

Introduction

Acute ischemic stroke (AIS) remains a leading cause of death and long-term disability worldwide and is associated with a high socioeconomic burden [1,2,3]. The current standard of care for AIS is thrombolysis by intravenous recombinant tissue plasminogen activator (rtPA, alteplase) within 4.5 h of stroke onset, and endovascular therapy in selected patients within 24 h of stroke onset [4,5,6,7,8]. Following AIS, activated (M1-phenotype) microglia produce pro-inflammatory mediators including tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 [19]. The latter are regulated by the mitogen-activated protein kinase kinase kinase (MAP3K) TGFβ-activated kinase 1 (TAK1). While activation of TAK1 is reported to exacerbate brain damage, pharmacological inhibition of TAK1 using 5Z-7Oxozeaenol exerts neuroprotection after subarachnoid hemorrhage as well as cerebral ischemia [23, 25, 26]

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