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Updated: Jan 18, 2026
Isolation and Flow Cytometric Assessment of Neuroimmune Interactions in a Mini-Stroke Murine Model
Published on: June 20, 2025
Miltirone attenuates post-ischemic stroke neuroinflammation and microglial lipid metabolism via regulating LBP and
1Department of Anesthesiology, The PLA Rocket Force Characteristic Medical Center, Beijing 100088, China.
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.
Insights
Miltirone, a compound from Salvia miltiorrhiza, offers neuroprotection against ischemic stroke by reducing inflammation, regulating lipid metabolism, and combating oxidative stress. This multi-target approach shows promise for clinical translation in stroke therapy.
Area of Science:
- Neuroscience
- Pharmacology
- Biochemistry
Background:
- Ischemic stroke is a primary cause of neurological disability with complex pathologies.
- Current treatments are insufficient for multifactorial stroke damage.
- Miltirone, from Salvia miltiorrhiza, exhibits antioxidant and anti-inflammatory properties, but its stroke-related neuroprotective mechanisms are unknown.
Purpose of the Study:
- To investigate the neuroprotective mechanisms of Miltirone in ischemic stroke.
- To evaluate Miltirone's effects on microglial polarization, lipid metabolism, and oxidative stress in stroke models.
- To elucidate the molecular pathways targeted by Miltirone in stroke.
Main Methods:
- Utilized young/aged distal Middle Cerebral Artery Occlusion (dMCAO) mouse models and oxygen-glucose deprivation/reperfusion (OGD/R) treated BV2 microglia.
- Assessed infarct volume, neurological function, microglial polarization (M1/M2), cytokine levels, lipid metabolism, oxidative stress markers (ROS, SOD, MDA), and the TLR4/MyD88/NF-κB pathway.
- Quantified outcomes using TTC staining, neurological tests, ELISA, immunofluorescence, RT-qPCR, Western blot, and flow cytometry.
Main Results:
- Miltirone significantly reduced infarct volume and brain edema, improving neurological and motor recovery in dMCAO mice.
- Miltirone promoted M2 microglial polarization, suppressed pro-inflammatory cytokines, and elevated anti-inflammatory cytokines.
- Miltirone restored lipid homeostasis, inhibited lipid synthesis, activated lipolysis, reduced oxidative stress, and suppressed the LBP/TLR4/MyD88/NF-κB pathway.
Conclusions:
- Miltirone demonstrates significant neuroprotection in ischemic stroke via multi-target actions.
- It effectively modulates neuroinflammation, lipid metabolism, and oxidative stress through the LBP/TLR4/NF-κB axis.
- Miltirone's comprehensive therapeutic potential makes it a promising candidate for clinical application in stroke treatment.
