Figure 6. 25-HC promoted oxygen-glucose deprivation/reoxygenation (OGD/R)-induced pyroptosis through activation of NLRP3 inflammasome in H9C2 cardiomyocytes. The H9C2cells were pretreated with NLRP3 inflammasome inhibitor BAY11-7082 (2 μM) 2 h before OGD/R treatment. Then, cells were treated with 25-hydroxycholesterol (0.4 μM) during reoxygenation. A, CCK-8 assay was used to detect cell viability. B, Lactate dehydrogenase (LDH) cytotoxicity detection assay was used to detect the LDH release. C, Hoechst 33342 and propidium iodide (PI) were used to stain cells; scale bars=100 μm. The nuclei were stained blue by Hoechst 33342, and the pyroptotic cells were stained red by PI. D, The protein expressions of NLRP3, Caspase-1, GSDMD, IL-1β, and GAPDH were detected by western blotting. Data are reported as means±SD (n=3). ***P<0.001 vs control group, ^##P<0.01 and ^###P<0.001 vs OGD/R group, ^&&P<0.01 and ^&&&P<0.001 vs OGD/R+25-HC group (one-way ANOVA with Tukey multiple comparison test).