益气养阴活血通络方对特发性肺纤维化模型大鼠血小板活化及肺组织CD40-CD40L系统的影响  

Effect of Yiqi Yangyin Huoxue Tongluo Formula(益气养阴活血通络方)on Platelet Activation and Lung Tissue CD40-CD40L System in Idiopathic Pulmonary Fibrosis Model Rats

作　　者：刘勇明[1] 李斯宇 庞立健[1] 臧凝子[1] 姜昕 王佳然 梁元钰 谢思梦 程美佳 闵冬雨[1] 吕晓东[2] LIU Yongming;LI Siyu;PANG Lijian;ZANG Ningzi;JIANG Xin;WANG Jiaran;LIANG Yuanyu;XIE Simeng;CHENG Meijia;MIN Dongyu;LYU Xiaodong(Affiliated Hospital of Liaoning University of Traditional Chinese medicine,Shenyang,110032;Liaoning University of Traditional Chinese medicine;Liaoning University of Traditional Chinese Medicine Hospital Subsidiary Fourth)

机构地区：[1]辽宁中医药大学附属医院,辽宁省沈阳市110032 [2]辽宁中医药大学 [3]辽宁中医药大学附属第四医院

出　　处：《中医杂志》2023年第5期504-511,共8页Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金(82204993,8227440,82004294);辽宁省教育厅重点攻关项目(L202067);辽宁省教育厅青年科技人才“育苗”项目(L202029,L202046);辽宁中医药大学自然科学类重点项目(2021LZY040)。

摘　　要：目的 基于CD40-CD40L系统探讨益气养阴活血通络方治疗特发性肺纤维化(IPF)的可能作用机制。方法 96只Wistar大鼠随机分为空白组、模型组、吡非尼酮组、益气养阴活血通络组,每组24只。除空白组外,其余各组大鼠采用气管内注射盐酸博来霉素制备IPF大鼠模型。造模次日吡非尼酮组采用吡非尼酮胶囊序贯方式灌胃,第1~7天给药剂量62.5 mg/(kg·d),第8~14天给药剂量125 mg/(kg·d),第15~28天给药剂量187.5 mg/(kg·d);益气养阴活血通络组给予益气养阴活血通络方颗粒剂18 g/(kg·d)灌胃;空白组和模型组每日给予10 ml/kg生理盐水灌胃。各组大鼠均每日灌胃1次,连续28天。分别于模型制备次日起第7、14、28天(各组每个时间点8只大鼠)时采用HE染色观察各组大鼠肺脏组织病理形态学变化;碱水解法检测肺组织羟脯氨酸(HYP)含量;酶联免疫吸附试验检测血清纤溶酶原激活物抑制因子(PAI)、凝血因子Ⅸ(FⅨ)、凝血因子Ⅶ(FⅦ)、血小板因子4 (PF-4)表达水平;流式细胞术检测血小板CD41、CD62P表达水平;蛋白免疫印迹法检测肺脏组织CD40、CD40L蛋白水平。结果 空白组大鼠肺组织病理HE染色显示正常。模型组大鼠病理显示第7天时肺泡间隔增宽、腔内炎症细胞聚集,肺间质可见明显水肿;第14天时肺间质内可见胶原纤维增生;第28天时肺泡逐渐萎缩,胶原纤维大量沉积。与空白组同时间点比较,模型组大鼠第7、14、28天肺组织HYP含量,血小板CD41、CD62P表达水平,血清PAI、FⅨ、FⅦ、PF-4表达水平,肺组织CD40、CD40L蛋白水平均显著升高(P<0.01)。与模型组同时间点比较,吡非尼酮组和益气养阴活血通络组大鼠肺组织病理形态学显示炎症细胞浸润明显改善、纤维化程度显著减轻。各给药组大鼠第7、14、28天肺组织HYP含量,血小板CD41表达水平,血清PAI、FⅨ、FⅦ、PF-4表达水平,肺组织CD40、CD40L蛋白水平均有不同程度下降(PObjectiveTo explore the possible mechanism of Yiqi Yangyin Huoxue Tongluo Formula(益气养阴活血通络方,YYHTF)in treating idiopathic pulmonary fibrosis(IPF)based on CD40-CD4OL system.Methods Totally,96 Wistar rats were randomly divided into blank group,model group,pirfenidone group,and YYHTF group,with 24 rats in each group.IPF rat model was established by intratracheal injection of bleomycin hydrochloride in all groups except for the blank group.On the next day after modeling,the pirfenidone group was given pirfenidone capsules by sequential gavage;the dosage was 62.5 mg/kg each day on the lst to 7th day,while 125 mg/kg on the 8th to 14th day,and 187.5 mg/kg on the 15th to 28th day.The YYHTF group was given YYHTF granules at the dosage of 18 g/kg by gavage.The blank group and model group were given 10 ml/kg of normal saline by gavage.All interventions were given once a day for 28 days.On the 7th,14th,and 28th day since the next day after modelling(8 rats were measured at each timepoint of each group),hematoxylin eosin(HE)staining was used to observe the pathological changes of lung tissue;the contents of hydroxyproline(HYP)in lung tissue of rats in each group were detected by alkaline hydrolysis,and the expression levels of plasminogen activator inhibitor(PAI),coagulation factor IX(FIX),coagulation factor VI(FVI)and platelet factor 4(PF-4)in serum were detected by enzyme-linked immunosorbent assay;the expression levels of CD41 and CD62P in platelets were detected by flow cytometry,and the protein levels of CD40 and CD4OL in lung tissue of rats were detected by western blotting.Results HE staining showed normal in the lung tissue of the rats in the blank group.In the model group,pathological results showed that the alveolar septum were widened,and the inflammatory cells in the lumen were gathered with obvious edema in the pulmonary interstitium on the 7th day;collagen fibers hyperplasia was seen in the pulmonary interstitium on the 14th day;the alveoli was gradually atrophied,and a large number of collagen

关 键 词：特发性肺纤维化 益气养阴活血通络方 血小板活化 CD40-CD40L系统 

分 类 号：R285.5[医药卫生—中药学]