Role Of CircHIPK2/miR-124/?-1R Axis In Astrocyte Activation

Objective:Most of the central nervous system(CNS)diseases are accompanied by severe neuroinflammatory disorders.Previous study has shown that neuroinflammatory disorders play important roles in CNS diseases.Astrocyte,the most abundant glial cell in mammalian brain,plays an integral role in neuroinflammation.However,the mechanisms of astrocyte activation are still poorly understood.This study shows the circular RNA HIPK2(circHIPK2)functions as an endogenous microRNA 124(miR-124)sponge and inhibits its activities to regulate astrocyte activation.These findings provide novel insights regarding the specific contribution of circHIPK2 to astrocyte activation in the context of drug abuse as well as for the treatment of a broad range of neuroinflammatory disorders.Methods:The expression of glial fibrillary acidic protein(GFAP),?-1R,endoplasmic reticulum(ER)stress associated proteins and autophagy associated proteins was detected by Western blot.The relative levels of circHIPK2,miR-124 and?-1R mRNA were detected by realtime PCR.Dual-luciferase reporter assay system was performed to demonstrate the interaction between miR-124 and ?-1R mRNA.The binding of circHIPK2 and miR-124 was demonstrated by RNA pull down assay and fluorescence in situ hybridization.The role of circHIPK2/miR-124/?-lR axis in astrocyte activation was illustrated by Western blot and immunostaining after lentivirus vector transfection in vitro and lentivirus vector microinjection in vivo.Results:1)?-lR participated in astrocyte activation induced by LPS;2)miR-124 participated in the upregulation of ?-1R induced by methylamphetamine;3)miR-124 inhibited astrocyte activation by targeting ?-1R in vitro;4)miR-124 inhibited astrocyte activation by targeting ?-1R in vivo;5)circHIPK2 binds miR-124;6)Knockdown of circHIPK2 expression inhibited astrocyte activation by targeting miR-124 in vitro;7)Knockdown of circHIPK2 expression inhibited astrocyte activation in the hippocampi of WT mice;8)Methylamphetamine treatment resulted in autophagy and ER stress in astrocyte;9)Knockdown of circHIPK2 expression inhibited astrocyte autophagy by targeting miR-124 in astrocyte;10)Knockdown of circHIPK2 expression inhibited ER stress in astrocyte by targeting miR-124 in astrocyte;11)ER stress promoted methamphetamine-induced astrocyte activation and autophagy inhibited astrocyte activation;12)ER stress induced by methamphetamine promoted autophagy,but autophagy inhibited ER stress induced by methamphetamine.Conclusions: CircHIPK2/miR-124/?-1R axis was involved in methylamphetamine/LPS-mediated astrocyte activation and regulated astrocyte activation via ER stress and autophagy...