基于NF-κB信号通路探究大黄酸对肺泡上皮细胞损伤的保护作用  

Protective effect of rhein on proliferation and migration of alveolar epithelial cells injured by lipopolysacchride and related mechanism

作　　者：赵宁生 胡勇[2] 郑玲 ZHAO Ningsheng;HU Yong;ZHENG Ling(Department of Respiratory and Critical Care Medicine,the First Affiliated Hospital of Guangxi University of Chinese Medicine,Nanning 530023,China;不详)

机构地区：[1]广西中医药大学第一附属医院呼吸与危重症医学科,南宁530023 [2]广西中医药大学第一附属医院中医经典科,南宁530023

出　　处：《浙江医学》2023年第12期1244-1248,1254,I0004,共7页Zhejiang Medical Journal

基　　金：广西中医药大学2022年研究生教育创新计划项目(2123210004)。

摘　　要：目的探讨大黄酸对脂多糖(LPS)诱导的炎症损伤人肺泡上皮细胞增殖、迁移及NF-κB信号通路的保护作用。方法体外培养A549细胞分为对照组(不做干预)、模型组(以10μg/ml LPS刺激A549细胞24 h构建体外急性肺损伤细胞模型)、LPS+2、4、8μmol/L大黄酸组(在模型组的基础上加2、4、8μmol/L大黄酸),采用ELISA法及细胞计数试剂盒-8(CCK-8)测定细胞炎症因子水平和活力以筛选出大黄酸最适浓度(2μmol/L)。而后将细胞分为对照组(不做干预)、模型组(10μg/ml LPS刺激A549细胞24 h)、大黄酸组(在模型组的基础上加2μmol/L大黄酸进行干预)、BAY 11-7082组(在模型组的基础上加5μmol/L BAY 11-7082进行干预)、抑制剂组(大黄酸组的基础上加5μmol/L BAY 11-7082进行干预)和激活剂组[大黄酸组的基础上加1μmol/L佛波酯(PMA)进行干预]。24 h后,分别采用5-乙炔基-2’脱氧尿嘧啶核苷(EdU)、Transwell小室及Western blot法测定各组细胞增殖率、迁移数及半胱氨酸天冬氨酸蛋白酶-1(Caspase-1)、细胞周期素D1(Cyclin D1)、NF-κB p65和磷酸化NF-κB p65(p-NF-κB p65)蛋白表达水平。结果与模型组相比,LPS+不同浓度大黄酸组炎症因子IL-6、IL-1β水平均降低(均P<0.05),细胞活力均升高(均P<0.05)。与对照组相比,模型组细胞增殖率、Cyclin D1蛋白表达水平均降低(均P<0.05),细胞迁移数、Caspase-1、p-NF-κB p65蛋白表达水平均升高(均P<0.05)。与模型组相比,大黄酸组和BAY 11-7082组显著扭转了上述指标的变化(均P<0.05)。与大黄酸组相比,BAY 11-7082加强了、PMA则削弱了2μmol/L大黄酸的作用(均P<0.05)。结论大黄酸对LPS诱导的炎症损伤A549细胞具有保护作用,并促进细胞增殖和抑制细胞迁移,其作用机制可能与抑制NF-κB信号通路有关。Objective To investigate the protective effect of rhein on proliferation,migration and NF-κB signaling pathway of human alveolar epithelial cells injured by lipopolysaccharide(LPS).Methods A549 cells cultured in vitro were divided into control group(no intervention),model group(A549 cells were stimulated by 10μg/ml LPS for 24 h to construct in vitro acute lung injury cell model),and 10μg/ml LPS+2,4,8μmol/L rhein group(2,4,8 were added on the basis of model group).The optimal concentration of rhein(2μmol/L)was determined by ELISA and cell counting kit 8(CCK-8).The cells were then divided into control group(no intervention),model group(10μg/ml LPS stimulated A549 cells for 24 h),rhein group(2μmol/L rhein was added to the model group for intervention),and BAY 11-7082 group(5μmol/L BAY 11-7082 was added to the model group for intervention),inhibitor group(5μmol/L BAY 11-7082 was added to the rhein group)and activator group(1μmol/L PMA was added to the rhein group for intervention).Twenty-four hours later,cell proliferation rate,migration number,the expression levels of Caspase-1,Cyclin D1,NF-κB p65 and phosphorylated NF-κB p65(p-NF-κB p65)protein were determined by 5-ethynyl-2'-deoxyuridine(EdU),Transwell chamber and Western blot assay.Results Compared with the model group,the levels of IL-6 and IL-1βin LPS+rhein groups with different concentrations were decreased(all P<0.05)and cell viability were increased(all P<0.05).Compared with the control group,cell proliferation rate and the expression level of Cyclin D1 protein in the model group were decreased(both P<0.05),number of migration cells,the expression levels of Caspase-1,p-NF-κB p65 protein were increased(all P<0.05).Compared with the model group,rhein group and BAY 11-7082 group significantly reversed the changes of the above indicators(all P<0.05).Compared with the rhein group,BAY 11-7082 enhanced and PMA weakened the effect of rhein(all P<0.05).Conclusion Rhein can protect A549 cells from inflammatory damage induced by LPS,promote cell prolifer

关 键 词：急性肺损伤 脂多糖 大黄酸 核因子-ΚB 增殖 

分 类 号：R285[医药卫生—中药学]