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Updated: Jun 22, 2025
Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke
Published on: January 3, 2025
Scutellarin alleviates microglia-mediated neuroinflammation and apoptosis after ischemic stroke through the
Zhaoda Duan1, Haolun Chen1, Wei Miao2
1Department of Anatomy and Histology/Embryology Faculty of Basic Medical Sciences Kunming Medical University Kunming China.
Abstract:
Microglia are resident immune cells in the central nervous system that are rapidly activated to mediate neuroinflammation and apoptosis, thereby aggravating brain tissue damage after ischemic stroke (IS). Although scutellarin has a specific therapeutic effect on IS, the potential target mechanism of its treatment has not been fully elucidated. In this study, we explored the potential mechanism of scutellarin in treating IS using network pharmacology. Lipopolysaccharide (LPS) was used to induce an in vitro BV-2 microglial cell model, while middle cerebral artery occlusion (MCAO) was used to induce an in vivo animal model. Our findings indicated that scutellarin promoted the recovery of cerebral blood flow in MCAO rats at 3 days, significantly different from that in the MCAO group. Western blotting and immunofluorescence revealed that scutellarin treatment of BV-2 microglial cells resulted in a significant reduction in the protein expression levels and incidence of cells immunopositive for p-NF-κB, TNF-α, IL-1β, Bax, and C-caspase-3. In contrast, the expression levels of p-PI3K, p-AKT, p-GSK3β, and Bcl-2 were further increased, significantly different from those in the LPS group. The PI3K inhibitor LY294002 had similar effects to scutellarin by inhibiting neuroinflammation and apoptosis in activated microglia. The results of the PI3K/AKT/GSK3β signaling pathway and NF-κB pathway in vivo in MCAO models induced microglia at 3 days were consistent with those obtained from in vitro cells. These findings indicate that scutellarin plays a neuroprotective role by reducing microglial neuroinflammation and apoptosis mediated by the activated PI3K/AKT/GSK3β/NF-κB signaling pathway.
Insights
Scutellarin protects the brain after ischemic stroke by reducing neuroinflammation and apoptosis in microglia. It modulates the PI3K/AKT/GSK3β/NF-κB signaling pathway, offering a potential therapeutic mechanism for stroke recovery.
Area of Science:
- Neuroscience
- Pharmacology
- Immunology
Background:
- Microglia are key immune cells in the central nervous system, exacerbating brain damage post-ischemic stroke (IS) through neuroinflammation and apoptosis.
- Scutellarin demonstrates therapeutic effects on IS, but its precise molecular targets remain unclear.
Purpose of the Study:
- To elucidate the underlying mechanism of scutellarin's therapeutic action in ischemic stroke using network pharmacology.
- To investigate scutellarin's effects on microglial activation, neuroinflammation, and apoptosis in both in vitro and in vivo models.
Main Methods:
- Established in vitro BV-2 microglial and in vivo middle cerebral artery occlusion (MCAO) rat models.
- Utilized Western blotting and immunofluorescence to assess protein expression levels.
- Investigated the involvement of PI3K/AKT/GSK3β and NF-κB signaling pathways.
Main Results:
- Scutellarin accelerated cerebral blood flow recovery in MCAO rats.
- In vitro, scutellarin reduced pro-inflammatory and apoptotic markers (p-NF-κB, TNF-α, IL-1β, Bax, C-caspase-3) while upregulating anti-apoptotic markers (p-PI3K, p-AKT, p-GSK3β, Bcl-2).
- In vivo results corroborated in vitro findings, confirming scutellarin's modulation of key signaling pathways.
Conclusions:
- Scutellarin exerts neuroprotection in ischemic stroke by inhibiting microglial neuroinflammation and apoptosis.
- The therapeutic effect is mediated through the modulation of the PI3K/AKT/GSK3β/NF-κB signaling pathway.
