二陈汤加味通过HMGB1/RAGE/NF-κB信号通路对COPD大鼠细支气管炎症的影响  

Effect of Modified Erchentang on Bronchioles of Rats with Chronic Obstructive Pulmonary Disease by HMGB1/RAGE/NF-κB Signaling Pathway

作　　者：尚立芝[1] 李耀洋 季书 谢文英[1] 尚皓梵 陈壮 刘高阳 王琦[1] SHANG Lizhi;LI Yaoyang;JI Shu;XIE Wenying;SHANG Haofan;CHEN Zhuang;LIU Gaoyang;WANG Qi(Henan University of Chinese Medicine,Zhengzhou 450046,China)

机构地区：[1]河南中医药大学,郑州450046

出　　处：《中国实验方剂学杂志》2023年第6期44-54,共11页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金面上项目(81573881);河南省科技攻关项目(182102311163);2021年河南中医药大学重点学科建设项目(15102040-2021-19);河南省研究生教育改革与质量提升工程项目(29104011-2)。

摘　　要：目的:观测二陈汤加味对慢性阻塞性肺疾病(COPD)大鼠细支气管中高迁移率族蛋白1(HMGB1)/晚期糖基化终产物受体(RAGE)/核转录因子-κB(NF-κB)通路中关键分子表达的影响,探讨二陈汤加味通过HMGB1/RAGE/NF-κB信号通路对COPD细支气管炎症的作用及分子机制。方法:60只SD大鼠随机分为6组:正常组、模型组、二陈汤加味低、中、高剂量组(5、10、20 g·kg^(-1))和丙酮酸乙酯(EP)组(HMGB1抑制剂),每组10只。以烟熏联合气管滴注脂多糖的方法制备COPD大鼠模型。二陈汤加味各干预组灌胃(ig)给药,并腹腔注射林格溶液4 mL;EP组ig等体积生理盐水,并腹腔注射EP(0.04 g·kg^(-1));正常组和模型组等体积生理盐水ig、并等体积林格溶液腹腔注射,连续干预21 d。酶联免疫吸附测定法检测支气管肺泡灌洗液(BALF)中HMGB1、趋化因子CXCL1和CXCL2、单核细胞趋化因子-1(MCP-1)的含量;实时荧光定量聚合酶链式反应(Realtime PCR)检测肺组织中HMGB1、RAGE和NF-κB p65 mRNA表达,免疫组化(IHC)检测细支气管组织中HMGB1、RAGE、磷酸化(p)-κB p65和α-平滑肌肌动蛋白(α-SMA)蛋白表达。结果:与正常组比较,模型组用力肺活量(FVC)、第1秒末时间肺活量(FEV1)和FEV1/FVC均显著降低(P<0.01);BALF中HMGB1、CXCL1、CXCL2和MCP-1的含量均显著升高(P<0.01);肺组织HMGB1、RAGE、NF-κB p65 mRNA表达显著增加(P<0.01);HMGB1、RAGE、p-NF-κB p65和α-SMA蛋白表达明显增强(P<0.05,P<0.01)。与模型组比较,二陈汤加味中、高剂量组FEV1/FVC均明显提高(P<0.05,P<0.01);BALF中HMGB1、CXCL1、CXCL2和MCP-1含量均明显降低(P<0.05,P<0.01);HMGB1、RAGE、NF-κB p65 mRNA表达明显减少(P<0.05,P<0.01);HMGB1、RAGE、p-NF-κB p65和α-SMA蛋白表达均明显减弱(P<0.05,P<0.01)。结论:二陈汤加味能有效对抗COPD细支气管炎症反应。其机制可能与通过下调HMGB1、RAGE mRNA表达,抑制NF-κB活化,减少HMGB1、CXCL1、CXCL2和MCP-1的释放,从而抑制细支�Objective:To study the effect of modified Erchentang on the expression of key molecules in the high mobility group Box 1 protein(HMGB1)/receptor for advanced glycation endproduct(RAGE)/nuclear factor-κB(NF-κB)signaling pathway in bronchioles of rats with chronic obstructive pulmonary disease(COPD),to explore the mechanism of modified Erchentang against bronchiolar inflammation of COPD rats via HMGB1/RAGE/NF-κB signaling pathway.Method:Sixty SD rats were randomly divided into normal group,model group,modified Erchentang low-,medium-and high-dose groups(5,10,20 g·kg^(-1)·d-1)and ethyl pyruvate(HMGB1 inhibitor)group,with 10 in each group.The COPD rat model was prepared by cigarette smoke combined with tracheal injection of lipopolysaccharide(LPS).After modeling,the modified Erchentang groups were given corresponding drugs(ig)and Ringer’s solution(4 mL,ip),while the EP group was treated with equal volume of normal saline(ig)and EP(0.04 g·kg^(-1)·d^(-1),ip).The normal group and the model group received equal volume of normal saline(ig)and Ringer’s solution(ip)for 21 consecutive days.The contents of HMGB1,chemokine(C-X-C motif)ligand 1(CXCL1),CXCL2 and monocyte chemotactic protein-1(MCP-1)in bronchoalveolar lavage fluid(BALF)were detected by enzyme-linked immunosorbent assay(ELISA).The mRNA expressions of HMGB1,RAGE and NF-κB p65 were determined by Real-time polymerase chain reaction(Real-time PCR),and the protein expressions of HMGB1,RAGE,p-NF-κB p65,and alpha-smooth muscle actin(α-SMA)in bronchioles tissue of rats were determined by immunohistochemistry(IHC).Result:Compared with the conditions in the normal group,the forced vital capacity(FVC),forced expiratory volume in the first second(FEV1)and FEV1/FVC in the model group were decreased(P<0.01)while the contents of HMGB1,CXCL1,CXCL2 and MCP-1 in BALF were increased(P<0.01).And the model group presented higher mRNA expressions of HMGB1,RAGE and NF-κB p65(P<0.01)and protein expressions of HMGB1,RAGE,p-NF-κB p65 andα-SMA(P<0.05,P<0.01)than the normal

关 键 词：慢性阻塞性肺疾病 二陈汤 高迁移率族蛋白1(HMGB1) 晚期糖基化终产物受体(RAGE) 核转录因子-κB(NF-κB) 

分 类 号：R2-0[医药卫生—中医学] R22R285.5R289R33