Mailuoning oral liquid alleviates permanent ischemic stroke by targeting CCR5 to modulate microglial polarization and PI3K/AKT/mTOR axis

Background Ischemic stroke (IS) triggers neuroinflammation cascades where microglial polarization is a pathological determinant. Mailuoning oral liquid (MLN O) is clinically utilized to prevent thrombosis and treat convalescent IS, but its pharmacological targets in permanent IS remain unclear.Aim of the study This study investigated whether MLN O alleviates permanent IS by targeting CCR5-mediated microglial polarization and PI3K/AKT/mTOR axis.Materials and methods A permanent middle cerebral artery occlusion (p-MCAO) rat model and LPS-stimulated BV2 microglial cells were employed. MLN O was administered both prophylactically (acute phase) and therapeutically (recovery phase). Neurological deficits, infarct volume, brain edema, histopathology, and coagulation function were assessed. Microglial phenotypes and morphology, CCL5/CCR5 axis, and PI3K/AKT/mTOR axis were analyzed using IF, IHC, Q-PCR, and Western blotting. The CCR5 antagonist maraviroc (MVC), exogenous CCL5, and Ccr5 siRNA were used for mechanistic validation. Network pharmacology analysis was employed to further confirmed the potential targets and signaling pathways of MLN O in IS treatment.Results Prophylactic MLN O significantly reduced infarct volume, brain edema, and neurological deficits, while improving coagulation function in p-MCAO rats. In both acute and recovery phases, MLN O promoted microglial polarization from the M1 (INOS, CD86) to M2 (CD206, Arg-1) phenotype, downregulated the CCL5/CCR5 axis, and inactivated the phosphorylation of PI3K/AKT/mTOR. In vitro, MLN O suppressed LPS-induced M1 polarization and suppressed NO production. Notably, MLN O and MVC showed comparable effects in vivo and in vitro on microglial polarization and PI3K/AKT/mTOR inactivation. Conversely, CCR5 knockdown by siRNA abolished the regulatory effect of MLN O on microglial polarization, and exogenous CCL5 reversed the therapeutic actions of MLN O, confirming CCR5 as a critical upstream mediator.Conclusions MLN O provides potential neuroprotection against permanent IS by targeting CCR5 to orchestrate microglial M1/M2 polarization and suppress PI3K/AKT/mTOR axis. These findings provide a robust pharmacological rationale for the clinical application of MLN O in preventing acute and treating convalescent permanent IS.