Genistein-3′-sodium Sulfonate Attenuates Cerebral Ischemia/reperfusion Injury in Rats by Down-regulating Microglial M1 Polarization Through α7nAChR-mediated Inhibition of NF-κb Signaling Pathway 

Abstract

Background: Microglial M1 depolarization mediated prolonged inflammation contributing to brain injury in ischemic stroke. Our previous study revealed that Genistein-3′-sodium sulfonate (GSS) exerted neuroprotective effects in ischemic stroke and reduced pro-inflammatory cytokines production. This study aimed to explore whether GSS protected against brain injury in ischemic stroke by regulating microglial M1 depolarization and its underlying mechanisms. Methods: We established transient middle cerebral artery occlusion and reperfusion (tMCAO) model in rats and used lipopolysaccharide (LPS)-stimulated BV2 microglial cells as in vitro model. Brain infarcted volume and neurological deficit were evaluated by TTC staining and Garcia assessment, respectively. M1-depolarized markers, α7nAChR and NF-κB signaling proteins were determined using western blot. Real-time PCR was used to determine the expression of M1 depolarization markers. Morphological changes, IL-1β expression and the nucleus translocation of P65-NF-κB were measured using immunofluorescent staining. The level of IL-1β was also determined using ELISA. Results: Our results showed that GSS treatment significantly reduced the brain infarcted volume and improved the neurological function in tMCAO rats. Meanwhile, GSS treatment also dramatically reduced microglia M1 depolarization, reversed α7nAChR expression, and inhibited the activation of NF-κB signaling in the ischemic penumbra brain regions. These effects of GSS were further verified in LPS-induced M1 depolarization of BV2 cells. Furthermore, pretreatment of α7nAChR inhibitor (α-BTX) significantly restrained the neuroprotective effect of GSS treatment in both tMCAO rats and LPS-stimulated BV2 cells. Alpha-BTX also blunted the regulating effects of GSS on GSM1 depolarization and α7nAChR expression.Conclusions: This study demonstrates that GSS protects against brain injury and neurological deficit in ischemic stroke by reducing microglia M1 depolarization to suppress neuroinflammation in peri-infarcted brain regions through upregulating α7nAChR and thereby inhibition of NF-κB signaling. Our findings uncover a potential molecular mechanism for GSS treatment in ischemic stroke.