Miltirone attenuates post-ischemic stroke neuroinflammation and microglial lipid metabolism via regulating LBP and TLR4/NF-κB Axis

Abstract

Background

Ischemic stroke is a leading cause of neurological disability. Current therapies fail to address its multifactorial pathologies. Miltirone, a bioactive compound from Salvia miltiorrhiza, has shown antioxidative and anti-inflammatory potential. However, its neuroprotective mechanisms in stroke remain unexplored.

Methods

Using young/aged dMCAO models and OGD/R-treated BV2 microglia, we evaluated Miltirone’s effects on infarct volume, neurological function, microglial polarization, lipid metabolism. Cerebral infarct volume was quantified by TTC staining. Neurological deficits were assessed via mNSS, rotarod, and adhesive removal tests. Cell viability was determined by CCK-8 assay. Pro-/anti-inflammatory cytokines, SOD activity and MDA content were measured by ELISA. Microglial polarization was analyzed via immunofluorescence and RT-qPCR. TLR4/MyD88/NF-κB pathway proteins and PLN2 were analyzed by Western blot. Lipid metabolism was evaluated by BODIPY staining. ROS was measured by flow cytometry

Results

Miltirone reduced cerebral infarct volume, attenuated brain edema, and improved neurological/motor recovery in dMCAO mice. It shifted microglial polarization toward the anti-inflammatory M2 phenotype by suppressing M1 markers and enhancing M2 markers. Miltirone downregulated pro-inflammatory cytokines while elevating anti-inflammatory cytokines. Miltirone restored lipid homeostasis by inhibiting lipid synthesis genes and activating lipolysis genes. This reduced lipid accumulation. Mechanistically, Miltirone suppressed LBP expression and TLR4/MyD88/NF-κB pathway. Moreover, Miltirone mitigated oxidative stress by lowering ROS, restoring SOD activity, and reducing lipid peroxidation.

Conclusion

Miltirone confers neuroprotection through multi-target actions. It simultaneously provides neuroinflammation, regulates lipid metabolism, and counters oxidative stress. This occurs via LBP/TLR4/NF-κB axis modulation. Its multitarget action addresses the complexity of ischemic stroke pathophysiology, positioning it as a promising therapeutic candidate for clinical translation.