LRRC8A contributes to neuroinflammation after acute ischemic stroke by interacting with S1PR1 regulating MAPK signal pathway

Abstract

Abstract Objective and design: Neuroinflammation exerts important roles in the progression of acute ischemic stroke (AIS). LRRC8A, an essential component of volume-regulated anion channel (VRAC) is expressed predominantly in central nervous system. The aim of this study is to investigate the role of LRRC8A in microglia-mediated neuroinflammation after AIS and the underlying mechanism. Materials and methods TTC assay, HE and Nissl staining, RT-PCR, ELISA assay, flow cytometry, Western blot (WB) and immunofluorescence were used to detect the effect of DCPIB on brain injury, neuroinflammation after AIS. Concomitantly, immunofluorescence, FLIM-FRET, and co-immunoprecipitation were used to examine the interplays between LRRC8A and S1P receptor-1(S1PR1). Results In mice tMCAO/R model, the administration of LRRC8A-dependent VRAC blocker DCPIB remarkably increased the survival rate and effectively reduced neuronal injury. In addition, DCPIB markedly reversed microglia polarization toward the M1 phenotype and enhanced M2 phenotype. More importantly, LRRC8A physically interacted with S1PR1 via C-terminal leucine-rich repeat domain (LRRD) and DCPIB weakened their interaction. Meanwhile, DCPIB also eliminated the activation MAPK pathway mediated by S1PR1. Conclusions The present study revealed a novel regulatory role of LRRC8A in regulating microglia-mediated neuroinflammation through interacting with S1PR1 by LRRD and elucidated a molecular mechanism for the effects of DCPIB on microglia polarization.