基于NF-κB信号通路探讨湿生扁蕾乙酸乙酯提取物对NCM460细胞凋亡的影响  

Effects of ethyl acetate extract of Gentianopsis paludosa on the apoptosis of NCM460 cells via NF-κB signaling pathway

作　　者：柳娜 王嘉鸣 徐婕 景明 LIU Na;WANG Jia-ming;XU Jie;JING Ming(College of Pharmacy,Gansu University of Chinese Medicine,Lanzhou 730000,China)

机构地区：[1]甘肃中医药大学药学院,甘肃兰州730000

出　　处：《中成药》2024年第10期3255-3262,共8页Chinese Traditional Patent Medicine

基　　金：国家自然科学基金资助项目(82160854);甘肃省高校产业支撑计划项目(2023CYZC-55)。

摘　　要：目的探讨湿生扁蕾乙酸乙酯提取物对人正常结肠上皮细胞NCM460凋亡的影响。方法采用10 g/mL脂多糖(LPS)诱导NCM460细胞建立炎症模型,给予湿生扁蕾乙酸乙酯提取物(50、100、200 g/mL)干预24 h,采用CCK-8试剂盒检测细胞存活率,ELISA法检测细胞中炎症因子IL-1β、IL-6水平,TUNEL染色技术检测细胞凋亡情况,Western blot法检测细胞凋亡相关蛋白Bcl-2、Bax、caspase-3、caspase-9及NF-κB信号通路相关蛋白NF-κB、p-NF-κB、IKKα/β、p-IKKα/β、IκBα、p-IκBα蛋白表达,免疫荧光技术观察NF-κB在细胞核中的表达。结果与对照组比较,LPS处理NCM460细胞24 h后细胞存活率降低(P<0.01),细胞凋亡加重(P<0.01),细胞中IL-1β、IL-6水平升高(P<0.01),Bax、caspase-3、caspase-9、p-IKKα/β/IKKα/β、p-IκBα/IκBα、p-NF-κB/NF-κB蛋白表达升高(P<0.01),Bcl-2蛋白表达降低(P<0.01),并促进NF-κB入核;与LPS组比较,湿生扁扁蕾乙酸乙酯提取物干预24 h后细胞存活率升高(P<0.05,P<0.01),细胞凋亡减轻(P<0.01),IL-1β、IL-6水平降低(P<0.01),Bax、caspase-3、caspase-9、p-IKKα/β/IKKα/β、p-IκBα/IκBα、p-NF-κB/NF-κB蛋白表达降低(P<0.05,P<0.01),Bcl-2蛋白表达升高(P<0.01),并抑制NF-κB入核。结论湿生扁蕾乙酸乙酯提取物可能通过调控NF-κB信号通路改善LPS诱导的NCM460细胞凋亡。AIM To investigate the effects of ethyl acetate extract of Gentianopsis paludosa(Hook.f.)Ma on the apoptosis of human normal colon epithelial cell NCM460.METHODS The NCM460 inflammatory cell model induced by 10 g/mL lipopolysaccharide(LPS)was administered with the ethyl acetate extract of G.paludosa(50,100,200 g/mL)for 24 hours.The cells had their survival rate detected by CCK-8 kit;their levels of inflammatory factors IL-1βand IL-6 detected by ELISA;their apoptosis detected by TUNEL staining;their expressions of apoptosis-related proteins Bcl-2,Bax,caspase-3,caspase-9 and NF-κB signaling pathway-related proteins NF-κB,p-NF-κB,IKKα/β,p-IKKα/β,IκBαand p-IκBαdetected by Western blot;and their nucleus NF-κB expression observed using the immunofluorescence technique.RESULTS Compared with the control group,the NCM460 cells exposed to LPS displayed decreased cell viability(P<0.01);increased cell apoptosis and levels of IL-1βand IL-6(P<0.01);decreased Bax,caspase-3,caspase-9,p-IKKα/β/IKKα/β,p-IκBα/IκBα,p-NF-κB/NF-κB protein expressions(P<0.01);and increased Bcl-2 protein expression(P<0.01).Compared with the model group,after 24 hours treatment of ethyl acetate extract of G.paludosa,the trial group displayed increased cell viability(P<0.05,P<0.01);decreased apoptosis(P<0.01);decreased levels of IL-1βand IL-6(P<0.01);decreased protein expressions of Bax,caspase-3,caspase-9,p-IKKα/β/IKKα/β,p-IκBα/IκBα,p-NF-κB/NF-κB(P<0.05,P<0.01);increased Bcl-2 protein expression(P<0.01);and inhibited NF-κB translocation into the nucleus.CONCLUSION Ethyl acetate extract of G.paludosa may improves the apoptosis of LPS-induced NCM460 cells via NF-κB signaling pathway.

关 键 词：湿生扁蕾 乙酸乙酯提取物 炎症性肠病 NCM460细胞 脂多糖 凋亡 NF-ΚB信号通路 

分 类 号：R285.5[医药卫生—中药学]