Reducing degradation of EETs by sEH protein macromolecule depletion prevents MCAO and inhibits microglia-mediated inflammatory response.

Endogenous endorphin (EETs) accumulation plays a neuroprotective role after cerebral ischemia-reperfusion injury, but the regulatory mechanism of its degradation remains unclear. With the deepening understanding of the role of sEH proteinin the metabolism of EETs, it has been suggested that depletion of sEH protein (encoded by EPHX2 gene) macromolecules may reduce the degradation of EETs, which is of great significance for improving cerebral ischemic injury. In this paper, we used CRISPR/Cas9 technique to construct EPHX2 gene knockout rat model and evaluate its role in MCAO. Primary microglia were isolated and transfected, cell viability analysis, ELISA, qPCR and immunohistochemistry were performed to systematically evaluate the effects of sEH on EETs degradation, neural function and inflammatory response. It was found that sEH depletion significantly alleviated the pathological changes and reduced the degradation of EETs in MCAO rats. Antagonistic EETs could reverse the neuroprotective effects of sEH depletion. Further analysis showed that sEH depletion inhibited the inflammatory response by regulating the phenotypic transition of microglia. Specific mechanisms include inhibition of IFN-γ-induced microglial inflammatory responses via EETs and inhibition of p-p38/NF-κB signaling pathways. Studies have revealed that sEH protein is a key regulator of EETs degradation, and sEH protein macromolecule depletion can effectively reduce the metabolic degradation of EETs, thereby alleviating cerebral ischemic injury and relieving inflammation.