Mcc950对H_2O_2诱导的ARPE-19细胞炎性损伤的保护作用  

Protection of Mcc950 against H_2O_2-induced inflammatory injury in ARPE-19 cells

作　　者：李淑艳 李晓峰[2] 许星照[3] 刘庆淮[1] Shu-Yan Li;Xiao-Feng Li;Xing-Zhao Xu;Qing-Huai Liu(Department of Ophthalmology,The First Affiliated Hospital with Nanjing Medical University,Nanjing 210029,Jiangsu Province,China;Department of Ophthalmology,The Affiliated Huai'an No.1 People's Hospital of Nanjing Medical University,Huai'an 223300,Jiangsu Province,China;Department of Ophthalmology,The First People's Hospital of Changzhou,Changzhou 213003,Jiangsu Province,China)

机构地区：[1]南京医科大学第一附属医院眼科,中国江苏省南京市210029 [2]南京医科大学附属淮安第一医院眼科,中国江苏省淮安市223300 [3]中国江苏省常州市第一人民医院眼科,213003

出　　处：《国际眼科杂志》2018年第9期1583-1588,共6页International Eye Science

摘　　要：目的:探讨Mcc950对过氧化氢(H_2O_2)诱导的人视网膜色素上皮细胞(ARPE-19细胞)炎性损伤的保护作用。方法:将ARPE-19细胞分为正常对照组、H_2O_2损伤组、单纯Mcc950给药组、Mcc950预处理+H_2O_2损伤组,采用CCK-8法检测细胞活力并确定H_2O_2和Mcc950合适的实验浓度,ELISA法检测细胞分泌的IL-1β浓度,免疫印迹(Western blot)法检测细胞中NLRP3炎性小体相关蛋白的表达情况,TUNEL染色法观察细胞凋亡情况。结果:细胞活力随H_2O_2刺激浓度的增加逐渐下降,H_2O_2浓度为400μmol/L时细胞活力显著降低;而0.1、1μmol/L Mcc950对细胞活力无显著影响,故选取400μmol/L H_2O_2和1μmol/L Mcc950作为合适的实验浓度。与正常对照组相比,H_2O_2损伤组细胞活力显著降低,细胞上清液中IL-1β浓度显著升高,细胞凋亡率明显增加,差异均有统计学意义(P<0.05),而与H_2O_2损伤组比较,Mcc950预处理+H_2O_2损伤组细胞活力明显升高,细胞上清液中IL-1β浓度和细胞凋亡率均降低,差异均有统计学意义(P<0.05)。Western blot检测结果显示,H_2O_2能够促进细胞中NLRP3、Pro-caspase1和caspase1的表达,而Mcc950预处理+H_2O_2损伤组细胞中NLRP3、Procaspase1依然保持高表达,但caspase1表达水平得到明显抑制,表明Mcc950能有效抑制NLRP3炎性小体的激活从而干扰具有细胞凋亡作用的成熟caspase1的产生。结论:Mcc950能够有效抑制H_2O_2诱导的NLRP3炎性小体激活,恢复细胞活力,抑制细胞凋亡。·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-1βwere 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-1βin 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-1βin 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.

关 键 词：Mcc950 NLRP3炎性小体 人视网膜色素上皮细胞 IL-1Β 年龄相关性黄斑变性 

分 类 号：R774.5[医药卫生—眼科]