苏黄止咳胶囊中白花前胡甲素对咳嗽变异性哮喘的改善作用  

Ameliorative effects of praeruptorin A from Suhuang antitussive capsules on cough variant asthma

作　　者：赵子瑶 江宏 欧永玉 陈孝源 吴楠 白子玉 张之昊 谭宁华[1] ZHAO Zi-yao;JIANG Hong;OU Yong-yu;CHEN Xiao-yuan;WU Nan;BAI Zi-yu;ZHANG Zhi-hao;TAN Ning-hua(School of Traditional Chinese Pharmacy,China Pharmaceutical University,Nanjing 211198,China;Beijing Haiyan Pharmaceutical Co.Ltd.,Yangtze River Pharmaceutical Group,Beijing 102206,China)

机构地区：[1]中国药科大学中药学院,江苏南京211198 [2]扬子江药业集团北京海燕药业有限公司,北京102206

出　　处：《中成药》2024年第9期2904-2914,共11页Chinese Traditional Patent Medicine

基　　金：国家自然科学基金项目(32070356)。

摘　　要：目的探讨苏黄止咳胶囊中白花前胡甲素对咳嗽变异性哮喘(CVA)的影响。方法将大鼠随机分为正常组、模型组、地塞米松组(0.5 mg/kg)、苏黄止咳胶囊组(7 g/kg)和白花前胡甲素低、中、高剂量组(15、30、60 mg/kg),采用腹腔注射致敏剂(1 mg/mL卵清蛋白和10 mg/mL氢氧化铝)和雾化吸入1%卵清蛋白的方法构建CVA大鼠模型,第14天起灌胃给予相应剂量药物,给药14 d后,HE、Masson和PAS染色观察肺组织病理变化,血细胞分析仪检测大鼠支气管肺泡灌洗液(BALF)中炎症细胞数,ELISA法检测BALF中IL-4、IL-5、IL-13、MUC5AC水平。采用脂多糖(LPS)诱导建立RAW264.7细胞炎症模型,分别以4、8、16μmol/L白花前胡甲素和0.25 mg/mL苏黄止咳胶囊处理,Griess法检测细胞中NO水平,荧光显微镜下观察细胞中ROS表达。RT-qPCR法检测肺组织和细胞中IL-6、IL-1β、COX-2、iNOS、PPAR-γmRNA表达,Western blot法检测肺组织和细胞中p-P65、P65、p-IκBα、IκBα、NLRP3、caspase-1(p20)、IL-1β蛋白表达。结果体内实验结果表明,白花前胡甲素能减少CVA大鼠咳嗽次数(P<0.01),延长咳嗽潜伏期(P<0.05,P<0.01),减少BALF中嗜酸性粒细胞和中性粒细胞数(P<0.05,P<0.01),降低BALF中IL-4、IL-5、IL-13、MUC5AC水平(P<0.01)和肺组织中IL-6、IL-1β、COX-2、iNOS mRNA表达(P<0.05,P<0.01),降低P65、IκBα蛋白磷酸化及NLRP3、caspase-1(p20)、IL-1β蛋白表达(P<0.05,P<0.01)。体外实验结果表明,白花前胡甲素能抑制LPS诱导的RAW264.7细胞中NO的释放,降低细胞中ROS水平(P<0.01);降低细胞中IL-1β、COX-2、iNOS mRNA表达(P<0.05,P<0.01),升高PPAR-γmRNA表达(P<0.05),降低P65、IκBα蛋白磷酸化和NLRP3蛋白表达(P<0.05,P<0.01)。结论白花前胡甲素可能是通过抑制NF-κB信号通路活化并降低NLRP3炎症小体的表达,发挥抗炎和止咳作用,从而改善咳嗽变异性哮喘,是苏黄止咳胶囊的主要止咳功效成分之一。AIM To explore the effects of praeruptorin A from Suhuang antitussive capsules on cough variant asthma(CVA).METHODS The rats were randomly divided into the normal group,the model group,the dexamethasone group(0.5 mg/kg),the Suhuang antitussive capsules group(7 g/kg)and the low,medium and high dose praeruptorin A groups(15,30 and 60 mg/kg).The rat model of CVA was established by intraperitoneal injection of sensitizer(1 mg/mL ovalbumin and 10 mg/mL aluminum hydroxide)and aerosol inhalation of 1%ovalbumin followed by the corresponding dosing of drugs by gavage initiated on the 14th day.Another 14 days later,the rats had their pathological pulmonary changes observed by HE,Masson and PAS stainings;their number of inflammatory cells in bronchoalveolar lavage fluid(BALF)detected by hematology analyzer;and their levels of IL-4,IL-5,IL-13 and MUC5AC in BALF detected by ELISA.The RAW264.7 cell inflammatory model induced by lipopolysaccharide(LPS)was treated with 4,8,16μmol/L praeruptorin A or 0.25 mg/mL Suhuang antitussive capsules,respectively.And the cells had their NO level detected by Griess method,and their ROS expression observed using fluorescence microscopy.The detections of the pulmonary and cellular mRNA expressions of IL-6,IL-1β,COX-2,iNOS and PPAR-γby RT-qPCR;and the protein expressions of p-P65,P65,p-IκBα,IκBα,NLRP3,caspase-1(p20)and IL-1βby Western blot were conducted in both the cells and the rats.RESULTS The in vivo result showed that praeruptorin A reduced the cough frequency(P<0.01);prolonged the cough latency(P<0.05,P<0.01);reduced the number of eosinophils and neutrophils in BALF(P<0.05,P<0.01);decreased the levels of IL-4,IL-5,IL-13 and MUC5AC in BALF and the pulmonary mRNA expressions of IL-6,IL-1β,COX-2 and iNOS(P<0.05,P<0.01);and decreased the phosphorylation of P65 and IκBαprotein and NLRP3,caspase-1(p20)and IL-1βprotein expressions(P<0.05,P<0.01)as well.The in vitro result showed that praeruptorin A inhibited the release of LPS-induced NO and reduce the ROS level(P<0.01);decreased the

关 键 词：白花前胡甲素 苏黄止咳胶囊 咳嗽变异性哮喘(CVA) NF-ΚB NLRP3 炎症 

分 类 号：R285.5[医药卫生—中药学]