Neuronal EphA4 Regulates OGD/R-Induced Apoptosis by Promoting Alternative Activation of Microglia

Area of Science:

• Neuroscience
• Cell Biology
• Molecular Biology

Background:

• Post-injury inflammation, driven by activated microglia, exacerbates neuropathology in ischemic injuries.
• Microglia exist in two states: classically activated (M1) and alternatively activated (M2).
• Shifting microglia from M1 to M2 dominance is a potential therapeutic strategy for ischemic injury.

Purpose of the Study:

• To investigate the role of neuronal erythropoietin-producing human hepatocellular carcinoma cell receptor 4 (EphA4) in ischemic brain injury.
• To determine if modulating EphA4/ephrin signaling impacts microglia polarization and subsequent neuropathology.
• To elucidate the underlying molecular mechanisms, including RhoA/ROCK2 signaling, involved in EphA4-mediated microglia regulation.

Main Methods:

• Utilized short hairpin RNA (shRNA) to knockdown neuronal EphA4 in an in vitro model of ischemic injury (oxygen-glucose deprivation and reperfusion - OGD/R).
• Assessed OGD/R-induced apoptosis and microglia proliferation.
• Analyzed microglia polarization towards M1 or M2 phenotypes and measured inflammatory mediators, anti-inflammatory cytokines, and neurotrophic factors.
• Investigated the involvement of RhoA/Rho-associated kinase 2 (ROCK2) signaling pathway.

Main Results:

• Neuronal EphA4 knockdown significantly attenuated OGD/R-induced apoptosis and microglia proliferation.
• Knockdown of EphA4 promoted a shift in microglia polarization towards the M2 phenotype.
• M2-polarized microglia released reduced pro-inflammatory mediators and increased anti-inflammatory cytokines and neurotrophic factors.
• The protective effects of EphA4 knockdown were mediated through the RhoA/ROCK2 signaling pathway.

Conclusions:

• Blockage of neuronal EphA4/ephrin signaling mitigates ischemic brain injury.
• Modulating EphA4 influences microglia polarization, favoring the beneficial M2 phenotype.
• The EphA4/ephrin pathway regulates microglia activation via the RhoA/ROCK2 signaling cascade, presenting a novel therapeutic target for ischemic stroke.