Functional mechanism of EGR3 in cerebral ischemia/reperfusion injury in rats by modulating transcription of pri-miR-146a/146b to miR-146 and suppressing SORT1 expression.

ABSTRACT

OBJECTIVE: EGR3 is implicated in angiogenesis in rats with cerebral ischemia/reperfusion injury (CIRI). This research aimed to explore the effect and in vivo and ex vivo mechanisms of EGR3 in CIRI. METHODS: CIRI rat models were established via middle cerebral artery occlusion. Cell models were established via oxygen-glucose deprivation/reoxygenation (OGD/R). Brain injury was assessed by neurological scoring, HE, and TTC staining. Inflammatory factors and oxidative stress markers were measured using corresponding kits. Mitochondrial membrane potential and mitochondrial respiration were examined by flow cytometry and respirometry. EGR3-miR-146 network was predicted on TransmiR v2.0 database. Target genes of miR-146 were screened on Starbase, Targetscan, and miRDB databases. miR-146 expression was determined by RT-qPCR. Levels of EGR3 and SORT1 were determined by Western blot. Binding relationships among EGR3, miR-146, and SORT1 were validated by dual-luciferase assay. EGR3, miR-146, and SORT1 levels were altered by injection or cell transfection to observe their functions. RESULTS: EGR3 was poorly-expressed in CIRI rats and OGD/R-induced neurons. EGR3 overexpression reduced inflammatory factor levels and attenuated oxidative stress and mitochondrial injury in CIRI rats and OGD/R-induced neurons. EGR3 bound to miR-146b promoter region. EGR3 promoted pri-miR-146a/146b processing and stimulated miR-146 transcription. miR-146 overexpression ameliorated oxidative stress and mitochondrial injury and miR-146 downregulation abolished the effect of EGR3 overexpression in vitro. miR-146 targeted SORT1. SORT1 overexpression invalidated the protective function of miR-146 overexpression on oxidative stress and mitochondrial injury in vitro. CONCLUSION: EGR3 protected against CIRI by mitigating oxidative stress and mitochondrial injury via the miR-146/SORT1 axis.