Abstract:AIM: To research the protection of Mcc950, the inhibitor of NLRP3, against the inflammatory injury to human retinal pigment epithelium cell line ARPE-19.
METHODS: Cultured cell line ARPE-19 was divided into control group, H2O2 treating group, Mcc950 treating group and Mcc950+H2O2 treating group. Different concentrations of H2O2 and Mcc950 were used to treat the cells. Cell activity was detected by using CKK8 and proper experimental concentration of H2O2 and Mcc950 were determined. After the treatment, the concentration of IL-1β were detected by using ELISA. The change of NLRP3 related proteins were detected by Western blot. And cell apoptosis was examined by TUNEL stain.
RESULTS: Cell ability was gradually decreased along with the increasing treating concentrations of H2O2. Cell ability showed statistical difference when the concentration of H2O2 arrived 400μmol/L. With the concentration of 0.1 and 1μmol/L, Mcc950 had no effect on cell ability. So we chose 400μmol/L H2O2 and 1μmol/L Mcc950 as the experimental concentrations. Compared with the normal control group, the cell viability in the H2O2 treating group was significantly reduced, the IL-1β in the supernatant was significantly increased, and the apoptosis rate was significantly increased, with statistically significant differences(P<0.05). In Mcc950+H2O2 treating group, cell viability was significantly increased, the IL-1β in the supernatant and the apoptosis rate were significantly decreased(P<0.05). By Western blot, after treated with 400μmol/L H2O2, the IL-1β, NLRP3, pro-caspase1 and caspase1 were obviously increased compared to control group. After treated with Mcc950, the NLRP3 and pro-caspase1 still were at high level, the expression of caspase1 was suppressed, which indicated that Mcc950 effectively inhibited the activation of NLRP3 inflammasome consequently disturbed the formation of caspase1.
CONCLUSION: Mcc950 can inhibits the function of NLRP3, leading to increasing of the cell ability and decreasing of the cell apoptosis.