Objectives: Accumulating data indicate that soluble Fas Ligand (sFasL) is involved in inflammatory cascades after cerebral ischemia. However, the potential role of sFasL in modulating the neuron-microglia crosstalk is barely understood. We hypothesized that sFasL triggers neuronal death and M1-microglia polarization after ischemic injury.
Methods: Middle cerebral artery occlusion (MCAO) was induced in both FasL mutant gld mice and wild-type C57BL/6J mice. Microglial polarization was determined by immunohistochemical staining and real-time polymerase chain reaction. Expression of sFasL was detected by western blot. The expression of sFasL in the medium after OGD was detected by ELISA assay. Activity of sFasL was blocked by using its neutralizing antibody. Neuronal survival and apoptosis were quantified.
Results: Expression of sFasL at protein level in the ischemic cortex was increased by 451.67% in wild-type stroke mice over the sham-operated brain. The gld mice demonstrated attenuated neuronal death and M1-microglial phenotype markers (iNOS and CD86) compared to wild-type mice after MCAO. Protein expression of sFasL was increased both in the neuronal and microglial culture medium after OGD. In addition, conditioned medium obtained from ischemic neuron drove microglial polarization toward the M1 phenotype producing more pro-inflammatory cytokines (IL-1β, TNF-α and MCP-1). Meanwhile, conditioned medium from ischemic microglia exacerbated neuronal death dramatically. sFasL neutralizing antibody, however, significantly decreased these changes. Furthermore, phosphorylated JAK2/STAT3(Y705 and S727) was increased in microglia after neuronal-conditioned medium treatment, which was decreased following inhibition of sFasL activity.
Conclusions: The gld mice demonstrated attenuated ischemic brain damage. sFasL is involved in modulating the crosstalk between these two cells in a contact-independent way. Microglia-released sFasL exacerbated neuronal death while neuron-released sFasL induced M1-microglia polarization after ischemic stroke. Neutralization of sFasL, thereby, may protect against ischemic brain damage and provide novel views on post-stroke neuroimmune regulation.