Role of microglial pyroptosis in hypoxic-ischemic brain damage.
- Author:
Lan-Lan TAN
1
;
Mei LI
;
Chen-Xi FENG
;
Li-Xiao XU
;
Xin DING
;
Bin SUN
;
Gen LI
;
Xing FENG
Author Information
1. Department of Neonatology, Children's Hospital of Soochow University, Suzhou, Jiangsu 215025, China. xing_feng66@suda.edu.cn.
- Publication Type:Journal Article
- MeSH:
Animals;
Brain/metabolism*;
Intracellular Signaling Peptides and Proteins;
Microglia/metabolism*;
Pyroptosis;
Rats
- From:
Chinese Journal of Contemporary Pediatrics
2020;22(11):1226-1232
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To investigate the role of microglial pyroptosis in hypoxic-ischemic brain damage.
METHODS:An oxygen-glucose deprivation/reoxygenation (OGD/R) model of rat microglial cells were cultured in vitro. Western blot was used to measure the expression of the pyroptosis-related proteins caspase-1, interleukin-1β (IL-1β), and N-terminal gasdermin D (GSDMD-N) at 0, 1, 3, 6, 12, and 24 hours after OGD/R. After the microglial cells were transfected with lentivirus-mediated silenced gasdermin D (GSDMD), immunofluorescence assay and Western blot were used to measure the transfection rate of GSDMD. Microglial cell lines were divided into three groups: normal control, negative control, and LV-sh_GSDMD (lentivirus-mediated GSDMD silencing). CCK-8 assay and LDH kit were used to observe the effect of GSDMD silencing on the viability and toxicity of microglial cells at 24 hours after OGD/R. Western blot was used to observe the effect of GSDMD silencing on the levels of caspase-1, GSDMD-N, and IL-1β in the microglial cells at 24 hours after OGD/R.
RESULTS:The expression levels of the pyroptosis-related proteins caspase-1, GSDMD-N, and IL-1β in microglial cells were upregulated since 0 hour after OGD/R and reached the peak levels at 24 hours. A microglial cell model of lentivirus-mediated GSDMD silencing was successfully constructed. At 24 hours after OGD/R, compared with the normal control group, the GSDMD silencing group had a significant increase in the cell viability and a significant reduction in the cytotoxicity (P<0.05), as well as significant reductions in the protein expression levels of caspase-1, GSDMD-N, and IL-1β in microglial cells (P<0.05).
CONCLUSIONS:Lentivirus silencing of the key substrate protein for pyroptosis GSDMD can alleviate hypoxic-ischemic brain damage, suggesting that microglial pyroptosis aggravates hypoxic-ischemic brain damage.
