Paeoniflorin Directly Targets ENO1 to Inhibit M1 Polarization of Microglia/Macrophages and Ameliorates EAE Disease

The chronic autoimmune disease multiple sclerosis (MS) now remains incurable. Paeoniflorin (PF), which is a monoterpene glucoside obtained from <i>Paeonia lactiflora Pall</i>, is recognized for neuroprotective and anti-inflammatory properties. However, the precise mechanism by which PF regulates MS is unclear. This work aims to elucidate the underlying mechanisms of PF in EAE, a well established animal model of MS, and to discover the target proteins that PF directly acts on. Our results revealed that PF administration can significantly attenuate the clinical symptoms of EAE and alleviate the central nervous system (CNS) inflammatory environment by inhibiting M1-type microglia/macrophages. Mechanistically, PF was found to directly interact with the glycolytic enzyme α-enolase (ENO1), inhibiting its enzymatic activity and expression to impair glucose metabolism, thereby suppressing microglia/macrophage M1 polarization and ameliorating CNS inflammation. Significantly, <i>Eno1</i> knockdown in microglia/macrophages diminished their pro-inflammatory phenotype, while treatment with ENOBlock or the specific knockout of <i>Eno1</i> in microglia led to EAE remission, underscoring the critical role of ENO1 in EAE progression. This study uncovers the molecular mechanism of PF in treating EAE, linking the anti-inflammatory property of PF to the glucose metabolism process, which will broaden the prospective applications of PF.