目的 探讨养阴清肺膏（YYQF）干预小鼠肺纤维化的作用及机制。方法 随机取12只小鼠作为对照组，其余动物经气管插管注入博来霉素（BLM）建立肺纤维化模型，造模后将小鼠随机分为模型组、吡非尼酮片(阳性药，270 mg·kg^-1)组和YYQF高、中、低剂量(18.0、9.0、4.5 mL·kg^-1)组，每组12只，连续给药28 d，末次给药后取材。计算小鼠肺脏指数，试剂盒法检测肺组织中羟脯氨酸（HYP）含量，HE染色和Masson染色观察肺组织病理改变和胶原沉积变化，酶联免疫吸附实验检测肺组织中肿瘤坏死因子（TNF-α）、白细胞介素-6（IL-6）的水平，免疫组化检测肺组织中平滑肌肌动蛋白（α-SMA）、胶原I（COL1A）的表达，Western blotting法检测肺组织中磷酸化磷脂酰肌醇3-激酶（PI3K）、蛋白激酶B（Akt）、哺乳动物雷帕霉素靶蛋白（m TOR）的蛋白含量。结果 与模型组比较，YYQF高剂量组肺脏指数和HYP含量显著降低（P<0.05、0.01）,TNF-α表达水平降低（P<0.01）,Akt、m TOR的蛋白含量减少（P<0.01）;YYQF中剂量组HYP含量降低（P<0.01）,TNF-α、IL-6表达水平降低（P<0.01、0.05）,α-SMA和COL1A的表达降低（P<0.05、0.01）,Akt、m TOR的蛋白含量减少（P<0.01）;YYQF低剂量组肺脏指数和HYP含量显著降低（P<0.05、0.01）,IL-6表达水平降低（P<0.05）,α-SMA和COL1A的表达降低（P<0.05、0.01）,PI3K、Akt、m TOR的蛋白含量减少（P<0.05、0.01）。病理检测结果显示，YYQF低剂量组模型小鼠肺组织结构较清晰，肺泡隔轻度增宽，纤维组织增生，肺脏内少量或局灶性分布的蓝染的胶原纤维。结论 YYQF可改善博来霉素诱导的小鼠肺纤维化，其作用机制可能与抑制PI3K/Akt/mTOR信号通路有关。

Objective To investigate the pharmacological effect and effector mechanism of Yangyin Qingfei Ointment（YYQF） in the intervention of pulmonary fibrosis in mice. Methods Twelve mice were randomly selected as the control group, while the remaining animals underwent tracheal intubation and injection of bleomycin（BLM） to establish a pulmonary fibrosis model. After model establishment, the mice were randomly divided into a model group, a pirfenidone tablet(270 mg·kg^-1) group, and YYQF high, medium, and low dose(18.0, 9.0, 4.5 mL·kg^-1) groups, with 12 mice in each group. The drugs were administered continuously for 28 days, and samples were taken after the last administration. The lung index of the mice was calculated, and the content of hydroxyproline（HYP） in lung tissue was detected using a reagent kit method. HE staining and Masson staining were used to observe the pathological changes and collagen deposition in lung tissue. Enzyme-linked immunosorbent assay（ELISA） was used to detect the levels of tumor necrosis factor（TNF-α） and interleukin-6（IL-6） in lung tissue. Immunohistochemistry was used to detect the expression of α-smooth muscle actin（α-SMA） and collagen I（COL1A） in lung tissue. Western blotting was used to detect the protein content of phosphorylated phosphatidylinositol 3-kinase（PI3K）, protein kinase B（Akt）, and mammalian target of rapamycin（mTOR） in lung tissue. Results Compared with the model group, the high-dose YYQF group exhibited significantly reduced lung index and HYP content（P < 0.05, 0.01）, decreased TNF-α expression level（P < 0.01）, and reduced protein content of Akt and mTOR（P < 0.01）. The medium-dose YYQF group showed reduced HYP content（P < 0.01）, decreased expression levels of TNF-α and IL-6（P < 0.01, 0.05）, reduced expression of α-SMA and COL1A（P < 0.05, 0.01）, and decreased protein content of Akt and mTOR（P < 0.01）. The low-dose YYQF group demonstrated significantly reduced lung index and HYP content（P < 0.05, 0.01）, decreased IL-6 expression level（P < 0.05）, reduced expression of α-SMA and COL1A（P < 0.05, 0.01）, and decreased protein content of PI3K, Akt, and mTOR（P < 0.05, 0.01）. Pathological examination results indicated that the lung tissue structure of the low-dose YYQF group model mice was relatively clear, with mildly widened alveolar septa, fibrous tissue proliferation, and a small or focal distribution of blue-stained collagen fibers within the lungs. Conclusion YYQF can improve bleomycin-induced pulmonary fibrosis, which may be related to the inhibition of PI3K/Akt/mTOR signaling pathway.

R965

国家科技重大专项(2018ZX09201-011)