基于代谢组学探讨金水尘肺方改善二氧化硅诱导大鼠矽肺肺纤维化的作用机制研究  

Metabolomics insights into the mechanism by which Jinshui Chenfei formula ameliorates silica-induced fibrosis in rats

作　　者：何汶芮 杨帆[1] 侯润苏 魏毓 赵虎雷 田燕歌[1] 李建生[1] 赵鹏 He Wenrui;Yang Fan;Hou Runsu;Wei Yu;Zhao Hulei;Tian Yange;Li Jiansheng;Zhao Peng(Co-Construction Collaborative Innovation Center for Chinese Medicine and Respiratory Diseases by Henan&Education Ministry of P.R.China/Henan Key Laboratory of Chinese Medicine for Respiratory Disease,Henan University of Chinese Medicine,Zhengzhou 450046,Henan,China)

机构地区：[1]河南中医药大学,呼吸疾病中医药防治省部共建协同创新中心/河南省中医药防治呼吸病重点实验室,河南郑州450046

出　　处：《中国中西医结合急救杂志》2023年第6期657-663,共7页Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care

基　　金：国家自然科学基金(82105048);河南省重点研发与推广专项(212102310356);河南省中医药科研专项课题(20-21ZYZD01)。

摘　　要：目的从内源性代谢物变化途径探讨金水尘肺方改善二氧化硅(SiO_(2))诱导的矽肺纤维化的作用机制.方法将32只SPF级雄性SD大鼠按随机数字表法分为正常对照组、模型组、金水尘肺方组(9.72 g·kg^(-1)·d^(-1))、汉防己甲素组(27 mg·kg^(-1)·d^(-1)).采用一次性非暴露式气管滴注SiO_(2)混悬液(250 mg/kg)的方法制备矽肺模型大鼠;制模后5~8周给予相应药物灌胃.第8周末,检测各组大鼠用力肺活量(FVC)、潮气量(TV)以及肺动态顺应性(Cydn)的变化;采用苏木素-伊红(HE)染色、马松(Masson)染色观察各组肺组织病理学改变,并采用Szapiel评分标准和Ashcroft评分标准评价病灶肺泡炎、纤维化程度;采用免疫组化法检测Ⅰ型胶原蛋白(COLⅠ)、Ⅲ型胶原蛋白(COLⅢ)的阳性染色;采用蛋白质免疫印迹试验(Western blotting)检测肺组织转化生长因子-β1(TGF-β1)、纤维连接蛋白(FN)、α-平滑肌肌动蛋白(α-SMA)的蛋白表达水平.进一步采用超高压液相色谱串联四级杆飞行时间质谱联用(UPLC-Q-TOF-MS)技术对大鼠血清样本进行差异代谢物筛选,并基于代谢分析网站MetaboAnalyst 5.0进行通路分析.结果与正常对照组比较,模型组大鼠肺组织出现明显病理学损伤,表现为肺泡结构完整性被破坏,纤维结节包裹SiO_(2)颗粒,肺泡炎评分和肺纤维化评分均明显升高[肺泡炎评分(分):2.62±0.27比0.20±0.15,肺纤维化评分(分):5.42±0.66比0.50±0.84,均P<0.01],肺功能指标Cydn、FVC和TV均明显降低[Cdyn(mL/cmH_(2)O):0.26±0.03比0.33±0.03,FVC(mL):8.09±0.47比10.99±0.38,TV(mL):1.95±0.19比2.53±0.26,均P<0.01];肺组织COLⅠ、COLⅢ阳性染色和α-SMA、FN、TGF-β1的蛋白表达水平均明显升高[COLⅠ(A值):13.47±1.76比5.77±0.45,COLⅢ(A值):10.39±0.47比6.19±0.77,FN蛋白表达(FN/GAPDH):0.33±0.02比0.21±0.07,α-SMA蛋白表达(α-SMA/GAPDH):1.78±0.16比1.11±0.24,TGF-β1蛋白表达(TGF-β1/GAPDH):0.52±0.10比0.11±0.46,均P<0.01].与�Objective To exploring the mechanism of Jinshui Chenfei formula(JCF)in ameliorating silica(SiO_(2))-induced silicosis fibrosis based on endogenous metabolite changes.Methods A total of 32 SPF male Sprague-Dawley(SD)rats were divided into normal control group,model group,JCF group(9.72 g kg^(-1)·d^(-1)),and Tetrandrine group(27 mg:kg^(-1)·d^(-1))according to random number table method.The experimental silicosis model was established by intratracheal injection with Si02 suspension(250 mg/kg)on day 1.From week 5-8,silicosis rats were treated with tetrandrine or JCF.On the end of week 8,the changes of pulmonary function index,including forced vital capacity(FVC),tidal volume(TV)and lung dynamic compliance(Cydn)were detected.The pathological changes of lung tissue were analyzed by hematoxyline-osin(HE)staining and Masson staining,the severity of focal alveolitis and fibrosis was also evaluated using the Szapiel scale and the Ashcroft scale,the positive staining of collagen I(COL I)and COLII was detected using immunohistochemistry;the protein expression of transforming growth factor-β1(TGF-β1),fibronectin(FN),andα-smooth muscle actin(α-SMA)were measured by Western blotting.The rat serum samples were further screened for differential metabolites using ultra performance liquid chromatographytandem quadrupole time of flight mass spectrometr(UPLC-Q-TOF-MS)and pathway analysis was performed based on MetaboAnalyst 5.0.Results Compared with those in the normal control group,pathological changes such as alveolar structure destruction,the fibrous nodules encapsulated SiO_(2)particles were increased in lung tissues of rats in model group,alveolitis score and pulmonary fibrosis score were significantly higher(alveolitis score:2.62±0.27 vs.0.20±0.15,pulmonary fibrosis score:5.42±0.66 vs.0.50±0.84,both P<0.01);pulmonary function index including Cydn,FVC,and TV were significantly decreased[Cdyn(mL/cmH_(2)O):0.26±0.03 vs.0.33±0.03,FVC(mL):8.09±0.47 vs.10.99±0.38,TV(mL):1.95±0.19 vs.2.53±0.26,all P<0.01];positive stain

关 键 词：金水尘肺方 矽肺 代谢组学 差异代谢物 

分 类 号：R285.5[医药卫生—中药学]