基于代谢组学研究中药定喘汤对中性粒细胞型哮喘小鼠的疗效及作用机制  

Study the mechanism of Dingchuan decoction in neutrophilic asthma mice on serum metabolomics

作　　者：陈柏林 李三[1,2] 朱黎明[1,2] 曾丹[1] Chen Bolin;Li San;Zhu Liming;Zeng Dan(Hunan Institute of Geriatrics,Hunan Provincial People′s Hospital(The First Affiliated Hospital of Hunan Normal University),Changsha 410016,China;Department of Respiratory and Critical Care Medicine,Hunan Provincial People′s Hospital(The First Affiliated Hospital of Hunan Normal University),Changsha 410016,China)

机构地区：[1]湖南省人民医院(湖南师范大学附属第一医院)湖南省老年医学研究所,长沙410016 [2]湖南省人民医院(湖南师范大学附属第一医院)呼吸与危重症医学科,长沙410016

出　　处：《中国医师杂志》2022年第11期1639-1648,共10页Journal of Chinese Physician

基　　金：湖南省卫生健康委科研项目(20201343);湖南省自然科学基金(2021JJ70097)。

摘　　要：目的基于代谢组学的方法, 从内源性代谢物的变化分析定喘汤对中性粒细胞型哮喘的干预效果及其可能机制。方法 40只C57BL/6小鼠随机分为5组, 分别为正常组(A组)、中性粒细胞型哮喘模型组(B组)、定喘汤低剂量治疗组(C组)、定喘汤中剂量治疗组(D组)、定喘汤高剂量治疗组(E组), 每组8只。B、C、D、E组采用卵清白蛋白(OVA)和完全弗氏佐剂(CFA)联合诱导致敏小鼠建立中性粒细胞型哮喘模型, C、D、E组采用不同生药浓度的定喘汤进行干预治疗。采用Buxco小动物肺功能检测仪评估小鼠的气道反应性;提取肺泡灌洗液(BALF)采用计数板计数其白细胞总数, 细胞涂片染色进行分类计数;HE染色观察肺组织的病理改变。采用高效液相色谱联用四级杆飞行时间质谱(UPLC-Q-TOF-MS/MS)检测各组小鼠的血清代谢物, 采用常用的主成分分析(PCA), 偏最小二乘判别分析(PLS-DA)、正交偏最小二乘判别分析(OPLS-DA)模型对血清中代谢物图谱进行统计分析, 从内源性代谢物的变化来反映定喘汤的干预效果及其可能机制。结果 (1)一般行为观察:除A组小鼠外, 其他各组小鼠在激发期间均出现不同程度的哮喘症状。定喘汤各治疗组(C、D、E组)小鼠症状较B组程度减轻。(2)气道反应性:B组小鼠对乙酰甲胆碱(MCh)的气道反应性随着吸入浓度的升高而增加, 各浓度MCh水平下的气道阻力均明显高于A组(均P<0.01);C、D、E组气道反应性低于B组(均P<0.01);D、E组小鼠气道反应性低于C组(均P<0.05);D、E组气道反应性比较差异无统计学意义(P>0.05)。(3)气道炎症细胞浸润:B组小鼠BALF中白细胞总数、中性粒细胞百分率明显高于A组(均P<0.01);C、D、E组白细胞总数、中性粒细胞百分率低于B组(均P<0.01);D、E组小鼠白细胞总数、中性粒细胞百分率低于C组(均P<0.01);D、E组之间比较差异无统计学意义(均P>0.05)。(4)肺组织病理改变:A组小鼠肺Objective Based on the method of metabonomics,the intervention effect of Dingchuan decoction on neutrophilic asthma and its possible mechanism were analyzed from the changes of endogenous metabolites.Methods Forty C57BL/6 mice were randomly divided into five groups:normal group(group A),neutrophilic asthma model group(group B),Dingchuan decoction low dose treatment group(group C),Dingchuan decoction medium dose treatment group(group D),and Dingchuan decoction high dose treatment group(group E),with 8 mice in each group.B/C/D/E group used ovalbumin(OVA)and complete Freund′s adjuvant(CFA)to induce sensitized mice to establish neutrophilic asthma model,and C/D/E group used Dingchuan decoction with different concentrations of crude drugs for intervention treatment.Buxco small animal lung function tester was used to evaluate the airway reactivity of mice;The total number of white blood cells in bronchoalveolar lavage fluid(BALF)was counted by counting plate,and the number was classified by cell smear staining;The pathological changes of lung tissue were observed by hematoxylin and eosin(HE)staining.The serum metabolites of mice in each group were detected by high performance liquid chromatography coupled with four pole time of flight mass spectrometry(UPLC-Q-TOF-MS/MS).The commonly used principal component analysis(PCA),partial least squares discriminant analysis(PLS-DA),orthogonal partial least squares discriminant analysis(OPLS-DA)models were used for statistical analysis of the metabolite profiles in serum.The intervention effect of Dingchuan decoction and its possible mechanism were reflected from the changes of endogenous metabolites.Results(1)General behavior observation:except mice in group A,mice in other groups showed asthma symptoms of different degrees during the challenge period.The symptoms of mice in each treatment group(group C,D,E)of Dingchuan decoction were less than those in group B.(2)Airway reactivity:the airway reactivity of mice in group B to methacholine(MCh)increased with the inhalation conce

关 键 词：定喘汤 中性粒细胞型哮喘 代谢组学 疾病模型 动物 

分 类 号：R285.5[医药卫生—中药学]