Erythropoietin Enhances Post-ischemic Migration and Phagocytosis and Alleviates the Activation of Inflammasomes in Human Microglial Cells.

Recombinant human erythropoietin (rhEPO) has been shown to exert anti-apoptotic and anti-inflammatory effects after cerebral ischemia. Inflammatory cytokines interleukin-1ß and -18 (IL-1ß and IL-18) are crucial mediators of apoptosis and are maturated by multiprotein complexes termed inflammasomes. Microglia are the first responders to post-ischemic brain damage and are a main source of inflammasomes. However, the impact of rhEPO on microglial activation and the subsequent induction of inflammasomes after ischemia remains elusive. To address this, we subjected human microglial clone 3 (HMC-3) cells to various durations of oxygen-glucose-deprivation/reperfusion (OGD/R) to assess the impact of rhEPO on cell viability, metabolic activity, oxidative stress, phagocytosis, migration, as well as on the regulation and activation of the NLRP1, NLRP3, NLRC4, and AIM2 inflammasomes. Administration of rhEPO mitigated OGD/R-induced oxidative stress and cell death. Additionally, it enhanced metabolic activity, migration and phagocytosis of HMC-3. Moreover, rhEPO attenuated post-ischemic activation and regulation of the NLRP1, NLRP3, NLRC4, and AIM2 inflammasomes as well as their downstream effectors CASPASE1 and IL-1ß. Pharmacological inhibition of NLRP3 via MCC950 had no effect on the activation of CASPASE1 and maturation of IL-1ß after OGD/R, but increased protein levels of NLRP1, NLRC4, and AIM2, suggesting compensatory activities among inflammasomes. We provide evidence that EPO-conveyed anti-inflammatory actions might be mediated via the regulation of the inflammasomes.

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

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-1ß 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.