Primary microglia culture Based on previously described protocols [37 (link)–39 (link)], primary microglial cells were obtained from the cerebral cortices of 3-day-old C57BL/6 mice. After removing the meninges, cortical tissue was digested using 0.05% trypsin. The separated cells were then cultured in Dulbecco's modified Eagle's medium-F12 supplemented with 10% fetal bovine serum, 5 ng/ml of Granulocyte–macrophage colony-stimulating factor (STEMCELL Technologies, 78017), and penicillin/streptomycin (100 U/ml and 100 mg/ml, respectively) at 37 °C in a 5% CO2 humidified incubator. After 10 days, the mixed cell population was dominated by astrocytes and formed a fused trophoblast. The microglia gradually proliferated and floated in the supernatant. Finally, the cells from the supernatant were harvested and seeded in 35-mm confocal dishes.
IHT for the primary microglial cells The cell cultures were placed in the same intermittent hypoxia chamber used for the experimental mice. The cells were exposed to 21% oxygen (8 min) and 8% oxygen (8 min) for 10 cycles.
IHT for the primary microglial cells The cell cultures were placed in the same intermittent hypoxia chamber used for the experimental mice. The cells were exposed to 21% oxygen (8 min) and 8% oxygen (8 min) for 10 cycles.
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