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Updated: Dec 2, 2025
Characterization and Isolation of Mouse Primary Microglia by Density Gradient Centrifugation
Published on: February 16, 2018
Sestrin2 regulates microglia polarization through mTOR-mediated autophagic flux to attenuate inflammation during
Tingting He1,2, Wanlu Li2, Yaying Song1,2
1Department of Neurology, Ruijin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200025, China.
Background:
Neuroinflammation is the major pathogenesis of cerebral ischemia. Microglia are activated and polarized to either the pro-inflammatory M1 phenotype or anti-inflammatory M2 phenotype, which act as a critical mediator of neuroinflammation. Sestrin2 has pro-survival properties against ischemic brain injury. However, whether sestrin2 has an anti-inflammatory function by shifting microglia polarization and its underlying mechanism is unknown.
Methods:
Adult male C57BL/6 mice (N = 108) underwent transient middle cerebral artery occlusion (tMCAO) and were treated with exogenous sestrin2. Neurological deficit scores and infarct volume were determined. Cell apoptosis was examined by TUNEL staining and Western blotting. The expression of inflammatory mediators, M1/M2-specific markers, and signaling pathways were detected by reverse transcription-polymerase chain reaction, immunostaining, and Western blotting. To explore the underlying mechanism, primary neurons were subjected to oxygen-glucose deprivation (OGD) and then treated with oxygenated condition medium of BV2 cells incubated with different doses of sestrin2.
Results:
Sestrin2 attenuated the neurological deficits, infarction volume, and cell apoptosis after tMCAO compared to those in the control (p < 0.05). Sestrin2 had an anti-inflammatory effect and could suppress M1 microglia polarization and promote M2 microglia polarization. Condition medium from BV2 cells cultured with sestrin2 reduced neuronal apoptosis after OGD in vitro. Furthermore, we demonstrated that sestrin2 drives microglia to the M2 phenotype by inhibiting the mammalian target of rapamycin (mTOR) signaling pathway and restoring autophagic flux.
Conclusions:
Sestrin2 exhibited neuroprotection by shifting microglia polarization from the M1 to M2 phenotype in ischemic mouse brain, which may be due to suppression of the mTOR signaling pathway and the restoration of autophagic flux.
Insights
Sestrin2 reduces brain damage after stroke by reprogramming inflammatory microglia to an anti-inflammatory state. This neuroprotective effect involves inhibiting mTOR signaling and restoring autophagy.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Neuroinflammation is central to cerebral ischemia pathogenesis.
- Microglia polarization (M1/M2) critically mediates neuroinflammation.
- Sestrin2 shows pro-survival effects in ischemic brain injury.
Purpose of the Study:
- To investigate sestrin2's anti-inflammatory function in microglia polarization.
- To elucidate the underlying molecular mechanisms of sestrin2's action.
Main Methods:
- Transient middle cerebral artery occlusion (tMCAO) model in mice.
- Assessment of neurological deficits, infarct volume, and apoptosis.
- Analysis of inflammatory mediators and M1/M2 markers.
- In vitro studies using oxygen-glucose deprivation (OGD) and BV2 cell conditioned medium.
Main Results:
- Sestrin2 treatment reduced neurological deficits, infarct volume, and apoptosis post-tMCAO.
- Sestrin2 suppressed M1 microglia polarization and promoted M2 polarization.
- Sestrin2 inhibited mTOR signaling and restored autophagic flux, driving M2 phenotype.
Conclusions:
- Sestrin2 confers neuroprotection in ischemic stroke by promoting M2 microglia polarization.
- This effect is mediated by suppressing mTOR signaling and restoring autophagic flux.
