基于血清代谢组学研究金水缓纤方治疗肺纤维化机制

Study on the mechanism of Jinshui Huanxian Formula for the treatment of pulmonary fibrosis based on serum metabolomics

目的:采用代谢组学技术探讨金水缓纤方治疗肺纤维化(PF)大鼠的机制。方法:将32只SD大鼠分为对照组、模型组、金水缓纤方组和吡非尼酮组,每组8只。建立PF大鼠模型,金水缓纤方组和吡非尼酮组于造模后第29—41天分别采用相应药物进行治疗。Masson染色观察大鼠肺组织病理变化,免疫组化法测定肺组织中Ⅰ型胶原(Col-Ⅰ)、Ⅲ型胶原(Col-Ⅲ)的表达。基于超高效液相色谱联合四级杆-静电场轨道阱高分辨质谱(UPLC Q-Exactive MS)系统,对大鼠血清进行非靶向代谢组学分析,筛选差异代谢物,探讨PF大鼠代谢网络的变化及金水缓纤方的调控作用。结果:模型组大鼠肺组织纤维化评分和Col-Ⅰ、Col-Ⅲ表达较对照组显著升高(P<0.01),金水缓纤方组纤维化评分和Col-Ⅰ、Col-Ⅲ表达均较模型组显著降低(P<0.01,P<0.05)。模型组有8种代谢物较对照组发生显著变化(FDR<0.2),其中二十碳五烯酸、18-羟基二十碳五烯酸、溶血磷脂酰胆碱18∶1和溶血磷脂酰胆碱20∶3显著下调(P<0.01,P<0.05),1-磷酸鞘氨醇和哌啶羧酸显著上调(P<0.01);金水缓纤方组有5种代谢物(溶血磷脂酰胆碱18∶1、1-磷酸鞘氨醇和十五烷酸除外)较模型组有回调趋势,其中二十碳五烯酸和溶血磷脂酰胆碱20∶3差异有统计学意义(P<0.05)。结论:金水缓纤方对PF大鼠胶原沉积有明显改善作用,其作用机制可能与其调节脂肪酸代谢、磷脂代谢和赖氨酸代谢有关。

Objective:To explore the mechanism of Jinshui Huanxian Formula for the treatment of rats with pulmonary fibrosis(PF)based on metabolomics.Methods:Thirty-two SD rats were divided into control group,model group,Jinshui Huanxian Formula group and pirfenidone group,with 8 rats in each group.After the establishment of rat model with PF,the Jinshui Huanxian Formula group and pirfenidone group were treated with corresponding drugs from day 29 to day 41.The pathological change of lung tissues was observed by Masson staining,and the expression of collagen I(Col-I)and collagen III(Col-III)in lung tissues was determined by immunohistochemistry.Based on UPLC Q-Exactive MS system,the serum of rats was analyzed by non-targeted metabolomics.Then differential metabolites were screened,so as to reveal the changes of the metabolic network in rats with PF and the regulatory effects of Jinshui Huanxian Formula.Results:The fibrosis scores and the expressions of Col-I and Col-III of lung tissues of rats in the model group were significantly increased compared with the control group(P<0.01),which were decreased in Jinshui Huanxian Formula group compared with the model group(P<0.01,P<0.05).There were 8 metabolites that changed significantly in the model group compared with the control group(FDR<0.2),among which eicosapentaenoic acid,18-hydroxyicosapentaenoic acid,lysophosphatidylcholine 18∶1 and lysophosphatidylcholine 20∶3 were significantly down-regulated(P<0.01,P<0.05),sphingosine-1-phosphate and pipecolic acid were significantly up-regulated(P<0.01).Compared with the model group,the levels of 5 metabolites(except lysophosphatidylcholine 18∶1,sphingosine-1-phosphate and pentadecanoic acid)in Jinshui Huanxian Formula group recovered,and eicosapentaenoic acid and lysophosphatidylcholine 20∶3 changed significantly(P<0.05).Conclusion:Jinshui Huanxian Formula could significantly improve collagen deposition in rats with PF,and it may exert the effects by regulating fatty acid metabolism,phospholipid metabolism and lysine metabolism.