Role of protein O-GlcNAc modification in oxidative stress injury in nerve cells of mice subjected to oxygen-glucose deprivation and restoration
10.3760/cma.j.cn131073.20210709.01127
- VernacularTitle:蛋白质O-GlcNAc修饰在氧糖剥夺/复氧复糖小鼠神经细胞氧化应激损伤中的作用
- Author:
Kefan WU
1
;
Aining ZHANG
;
Zhen QIU
;
Meng JIANG
;
Zhongyuan XIA
Author Information
1. 武汉大学人民医院麻醉科 430060
- Keywords:
Glycosylation;
Reperfusion injury;
Brain;
Oxidative stress
- From:
Chinese Journal of Anesthesiology
2021;41(11):1396-1399
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To evaluate the role of protein O-linked beta-N-acetylglucosaminylation (O-GlcNAcylation) modification in oxidative stress injury in nerve cells of mice subjected to oxygen-glucose deprivation and restoration (OGD/R).Methods:The standard mouse hippocampal neuron cell line was inoculated on a culture plate or dish at a density of 5×10 4 cells/ml and divided into 4 groups ( n=20 each) using a random number table method: normal group (N group), O-(connection)N-acetylglucosamine hydrolase (OGA) inhibitor Thiamet G group (T group), OGD/R group (D/R group) and Thiamet G+ OGD/R complex sugar group (T-D/R group). The cells were exposed to a mixed gas of 94% N 2-5% CO 2-1% O 2 for 6 h in a low-glucose medium, then medium was replaced with a common medium for restoring oxygen and glucose, and the cells were cultured for 12 h. Thiamet G at a final concentration of 1 mmol/L was added to the culture medium at 4 h before OGD/R in T-D/R group, and the medium was replaced with a medium containing Thiamet G at a final concentration of 1 mmol/L at 4 h before extraction of cellular proteins.After oxygen and glucose restoration was completed, the accumulation of cellular ROS was measured using DCFH-DA staining, mitochondrial membrane potential was measured using Jc-1 staining, O-GlcNAc modification was determined by immunofluorescence, and the expression of nuclear factor E2-related factor 2 (Nrf2), c-Jun N-terminal kinase (JNK), phosphorylated JNK (p-JNK), and p53 tumor suppressor gene (p53) was detected using Western blot. Results:Compared with group N, the expression of O-GlcNAc in nerve cells was significantly up-regulated in group T, and the accumulation of ROS in nerve cells was significantly increased, JC-1 monomer was increased, JC-1 polymer was decreased, Nrf2 expression was down-regulated, and the expression of p-JNK and p53 was up-regulated in group D/R, and the expression of O-GlcNAc in nerve cells was up-regulated, the accumulation of ROS was increased, the polymerization of JC-1 monomer and JC-1 was increased, Nrf2 expression was down-regulated, and the expression of p-JNK and p53 was up-regulated in group T-D/R ( P<0.05). Compared with group D/R, the expression of O-GlcNAc in nerve cells was significantly up-regulated, the accumulation of ROS was decreased, JC-1 monomer was decreased, JC-1 polymer was increased, the expression of Nrf2 was up-regulated, and the expression of p-JNK and p53 was down-regulated in group T-D/R ( P<0.05). Conclusion:When mouse nerve cells are subjected to OGD/R, the protein O-GlcNAc modification as an endogenous protective mechanism is enhanced, which can reduce oxidative stress injury, and the mechanism may be related to regulating the Nrf2-mediated JNK pathway.
