RESUMO
Objective:To evaluate the role of silent information regulator-1 (SIRT1)/nucleotide-binding domain (NOD)-like receptor protein-3 (NLRP3) signaling pathway in sevoflurane postconditioning-induced attenuation of oxygen-glucose deprivation and restoration (OGD/R) injury in mouse hippocampal neuronal cell line (HT22) cells.Methods:The HT22 cells were seeded in a culture plate (96-well plate, 100 μl/well; 6-well plate, 2 ml/well) at the density of 5×10 4 cells/ml or in a culture dish (6 cm in diameter) and then divided into 4 groups ( n=24 each) using a random number table method: control group (Control group), OGD/R group, sevoflurane postconditioning group (SPC group), and SIRT1 small interfering RNA group (si-SIRT 1 group). In Control group, cells were cultured at 37 ℃ in normal culture atmosphere. In OGD/R group, the culture medium was replaced with glucose-free serum-free culture medium, and cells were exposed to 95% N 2+ 5% CO 2 for 4 h in an incubator at 37 ℃, and then the glucose-free serum-free culture medium was replaced with the primary culture medium, and cells were cultured for 24 h at 37 ℃ in normal culture atmosphere. In SPC group, the glucose-free serum-free culture medium was replaced with the primary cell culture medium after 4-h oxygen and glucose deprivation, the cells were put into the hypoxia incubator chamber which was filled with 2% sevoflurane immediately after start of reoxygenation, then the chamber was placed in an incubator and the cells were cultured for 1 h at 37 ℃ in normal culture atmosphere, and finally the cells were removed from the chamber and cultured for 23 h at 37 ℃ in normal culture atmosphere. In si-SIRT1 group, SIRT1 small interfering RNA 150 pmol was added at 24 h before surgery, cells were then incubated, and the other procedures were the same as those previously described in group SPC. The cell survival rate was determined using MTT assay. TUNEL assay was used to detect cell apoptosis, and the apoptosis rate was calculated. The expression of SIRT1, NLRP3, IL-1β and IL-18 mRNA was determined using polymerase chain reaction. The expression of SIRT1, NLRP3, interleukin-1beta (IL-1β) and IL-18 was detected using Western blot. Results:Compared with Control group, the cell survival rate was significantly decreased, the apoptosis rate was increased, the expression of SIRT1 protein and mRNA was down-regulated, and the expression of NLRP3, IL-1β and IL-18 protein and mRNA was up-regulated in OGD/R group ( P<0.05). Compared with OGD/R group, the cell survival rate was significantly increased, the apoptosis rate was decreased, the expression of SIRT1 protein and mRNA was up-regulated, and the expression of NLRP3, IL-1β and IL-18 protein and mRNA was down-regulated in SPC group ( P<0.05). Compared with SPC group, the cell survival rate was significantly decreased, the apoptosis rate was increased, the expression of SIRT1 protein and mRNA was down-regulated, and the expression of NLRP3, IL-1β and IL-18 protein and mRNA was up-regulated in si-SIRT1 group ( P<0.05). Conclusions:Activation of SIRT1-NLRP3 signaling pathway is involved in sevoflurane postconditioning-induced attenuation of OGD/R injury in HT22 cells.