Background: Activated microglia play a key role in initiating the inflammatory cascade following ischemic stroke and exert proinflammatory or anti-inflammatory effects, depending on whether they are polarized toward the M1 or M2 phenotype. The present study investigated the regulatory effect of icaritin (ICT) on microglial polarization in rats after cerebral ischemia/reperfusion injury (CI/RI) and explored the possible anti-inflammatory mechanisms of ICT.
Methods: A rat model of transient middle cerebral artery occlusion (tMCAO) was established. Following treatment with ICT, a G protein-coupled estrogen receptor (GPER) inhibitor or an extracellular signal-regulated kinase (ERK) inhibitor, the Garcia scale and rotarod test were used to assess neurological and locomotor function. 2,3,5-Triphenyltetrazolium chloride (TTC) and Fluoro-Jade C (FJC) staining were used to evaluate the infarct volume and neuronal death. The levels of inflammatory factors in the ischemic penumbra were evaluated using enzyme-linked immunosorbent assays (ELISAs). In addition, western blotting, immunofluorescence staining and quantitative PCR (qPCR) were performed to measure the expression levels of markers of different microglial phenotypes and proteins related to the GPER-ERK-nuclear factor kappa B (NF-κB) signaling pathway.
Results: ICT treatment significantly decreased the cerebral infarct volume, brain water content and fluorescence intensity of FJC; improved the Garcia score; increased the latency to fall and rotation speed in the rotarod test; decreased the levels of interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), Iba1, CD40, CD68 and p-P65-NF-κB; and increased the levels of CD206 and p-ERK. U0126 (an inhibitor of ERK) and G15 (a selective antagonist of GPER) antagonized these effects.
Conclusions: These findings indicate that ICT plays roles in inhibiting the inflammatory response and achieving neuroprotection by regulating GPER-ERK-NF-κB signaling and then inhibiting microglial activation and M1 polarization while promoting M2 polarization, which provides a new therapeutic for against cerebral ischemic stroke.
Keywords: Cerebral ischemia/reperfusion injury; ERK; GPER; Icaritin; Microglia; NF-κB.
© 2022. The Author(s).