Research Paper Volume 15, Issue 10 pp 4429—4443

Figure 2. Metformin inhibits inflammation and KGN cell pyroptosis by decreasing ROS production and oxidative stress. (A–E) Cells were pretreated with or without 10 μM LPS for 24 h and then incubated with 20 μM metformin for another 12 h. Intracellular levels of ROS, MDA, SOD, GSH, and GSH-px were measured using respective commercial kits; (F–J) Cells were pretreated with 10 μM LPS for 24 h and then incubated with 20 μM metformin and 10 mM NAC for another 12 h; (F) RT-PCR detected the mRNA levels of NLRP3, caspase-1, ASC, and GSDMD; (G) The protein levels of NLRP3, pro-caspase-1, cleaved-caspase-1, ASC and GSDMD-N were detected by Western blot; (H) Levels of LDH in the cell culture medium were examined using an LDH cytotoxicity detection kit; (I) Caspase-1 activity was assessed using a colorimetric caspase-1 activity assay kit; (J) The levels of IL-1β, IL-18, IL-6, and TNF-α in the cell culture supernatant were determined by ELISA. Data are represented as the means ± SD from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns. not significant.