Inhibition of Toll-like receptor 9 activation in microglia after oxygen-glucose deprivation and reoxygenation protects neurons from damage

AIM: To observe the Toll-like receptor 9 (TLR9) activation in microglia BV-2 cells after oxygen-glucose deprivation and reoxygenation (OGDR), and its effects on neuronal apoptosis. METHODS: The BV-2 cell supernatants were collected after the corresponding treatment and added to mouse primary cortical neurons after OGDR for 4 h, followed by normal culture for 24 h. The cells were divided into normal BV-2 group, NC-siRNA group, TLR9-siRNA group, OGDR group, OGDR+NC-siRNA group, OGDR+TLR9-siRNA group and control group (without adding BV-2 cell supernatant). The changes of the neuronal morphology were observed under an inverted phase- contrast microscope, and the neuronal apoptosis was detected by TUNEL. The protein expression of cleaved caspase-3 was detected by Western blotting. RESULTS: After OGDR, the axon turned thin, twisted and broken, and neuronal swelling, decrease in refraction and vacuolar degeneration were observed. The green-stained apoptotic bodies in the neurons in all groups were positive. Compared with control group, the caspase-3 protein levels in other groups were increased. Compared with the normal BV-2 group, the caspase-3 protein in OGDR group and TLR9-siRNA group was increased. Compared with OGDR+TLR9-siRNA group, the caspase-3 protein in TLR9-siRNA group and OGDR group was decreased. CONCLUSION: After OGDR, TLR9 activation in BV-2 cells induces neuronal apoptosis with the increase in caspase-3 protein level. Inhibition of TLR9 expression reduces neuronal damage.