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Characterization and Isolation of Mouse Primary Microglia by Density Gradient Centrifugation
Published on: February 16, 2018
Hydrogen inhibits microglial activation and regulates microglial phenotype in a mouse middle cerebral artery
Jun-Long Huang1, Wen-Wu Liu2, Anatol Manaenko3
1Discipline of Neuroscience, Department of Anatomy, Histology and Embryology, Shanghai Jiao Tong University School of Medicine; Department of Navy Aviation Medicine, Faculty of Naval Medicine, the Naval Military Medical University; Department of Navy Aviation Medicine, Naval Medical center of PLA, the Naval Military Medical University, Shanghai, China.
Abstract:
Microglia participate in bi-directional control of brain repair after stroke. Previous studies have demonstrated that hydrogen protects brain after ischemia/reperfusion (I/R) by inhibiting inflammation, but the specific mechanism of anti-inflammatory effect of hydrogen is poorly understood. The goal of our study is to investigate whether inhalation of high concentration hydrogen (HCH) is able to attenuate I/R-induced microglia activation. Eighty C57B/L male mice were divided into four groups: sham, I/R, I/R + HCH and I/R + N2/O2 groups. Assessment of animals happened in "blind" matter. I/R was induced by occlusion of middle cerebral artery for one hour). After one hour, filament was withdrawn, which induced reperfusion. Hydrogen treated I/R animals inhaled mix of 66.7% H2 balanced with O2 for 90 minutes, starting immediately after initiation of reperfusion. Control animals (N2/O2) inhaled mix in which hydrogen was replaced with N2 for the same time (90 minutes). The brain injury, such as brain infarction and development of brain edema, as well as neurobehavioral deficits were determined 23 hours after reperfusion. Effect of HCH on microglia activation in the ischemic penumbra was investigated by immunostaining also 23 hours after reperfusion. mRNA expression of inflammation related genes was detected by PCR. Our results showed that HCH attenuated brain injury and consequently reduced neurological dysfunction after I/R. Furthermore, we demonstrated that HCH directed microglia polarization towards anti-inflammatory M2 polarization. This study indicates hydrogen may exert neuroprotective effects by inhibiting the microglial activation and regulating microglial polarization. This study was conducted in agreement with the Animal Care and Use Committee (IACUC) and Institutional Animal Care guidelines regulation (Shanghai Jiao Tong University, China (approval No. A2015-011) in November 2015.
Insights
High concentration hydrogen inhalation reduces brain injury after stroke by modulating microglia activation. This neuroprotective effect involves shifting microglia towards an anti-inflammatory M2 state.
Area of Science:
- Neuroscience
- Immunology
- Biomedical Engineering
Background:
- Microglia play a dual role in brain repair post-stroke.
- Hydrogen shows promise in protecting the brain from ischemia/reperfusion (I/R) injury by reducing inflammation.
- The precise anti-inflammatory mechanisms of hydrogen, particularly its effect on microglia, require further elucidation.
Purpose of the Study:
- To investigate if high-concentration hydrogen (HCH) inhalation can mitigate I/R-induced microglia activation.
- To explore the neuroprotective effects of HCH on brain injury and neurological deficits following stroke.
- To determine HCH's impact on microglia polarization in the ischemic penumbra.
Main Methods:
- Male C57B/L mice underwent middle cerebral artery occlusion to induce I/R injury.
- Animals were divided into sham, I/R, I/R + HCH, and I/R + N2/O2 groups.
- HCH (66.7% H2/O2) or N2/O2 was inhaled for 90 minutes post-reperfusion; brain injury, neurobehavior, microglia activation, and gene expression were assessed.
Main Results:
- HCH inhalation significantly attenuated I/R-induced brain injury, including infarction and edema.
- Neurobehavioral deficits were reduced in mice treated with HCH.
- HCH treatment promoted microglia polarization towards the anti-inflammatory M2 phenotype in the ischemic penumbra.
Conclusions:
- High-concentration hydrogen inhalation exerts neuroprotective effects against I/R injury.
- Hydrogen's mechanism involves the inhibition of microglial activation and the regulation of microglia polarization to an M2 phenotype.
- These findings suggest hydrogen as a potential therapeutic agent for stroke recovery.
