虎杖苷对慢性阻塞性肺疾病大鼠气道炎症及TLR4/NF-κB信号通路的影响
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摘要
目的:探究虎杖苷对慢性阻塞性肺疾病(COPD)大鼠气道炎症的缓减作用,并初步探究其作用机制。方法:将大鼠随机分为空白对照组、模型组、虎杖苷30、60、120mg/kg组、地塞米松0.09mg/kg组。采用肺功能检测仪检测用力肺活量(FVC)、第0.1秒用力呼气容积(FEV0.1)、呼气峰流速(PEF)、最大呼吸中段气流(MMF)、肺顺应性(Cldyn);吉姆萨染色法检测支气管肺泡灌洗液(BALF)中白细胞总数及中性粒细胞(PMN)、肺泡巨噬细胞(AM)、淋巴细胞(LYM)比例;酶联免疫吸附法检测BALF中TNF-α、IL-8、IL-1β水平;HE染色观察肺组织病理学变化;高碘酸希夫氏染色(PAS)观察支气管病理学变化;免疫印迹法检测肺组织中TLR4、NF-κB、基质金属蛋白酶2(MMP-2)、MMP-9蛋白表达;免疫组化法检测肺组织中TLR4、NF-κB蛋白阳性表达率。结果:与对照组相比,模型组肺组织病理半定量评分显著升高;FVC、FEV0.1、PEF、MMF、Cldyn明显降低,BALF中AM细胞比例明显降低,白细胞总数、PMN、LYM细胞比例明显升高,TNF-α、IL-8、IL-1β水平明显升高,支气管气管壁、平滑肌厚度升高,肺组织中TLR4、NF-κB p65、MMP-2、MMP-9蛋白表达及TLR4、NF-κB p65蛋白阳性表达率明显升高。与模型组相比,虎杖苷各组、地塞米松组肺组织病理半定量评分显著降低。FVC、FEV0.1、PEF、MMF、Cldyn升高,BALF中AM细胞比例显著升高,白细胞总数、PMN、LYM细胞比例显著降低,TNF-α、IL-8、IL-1β水平显著降低,支气管气管壁、平滑肌厚度降低,肺组织中TLR4、NF-κB p65、MMP-2、MMP-9蛋白表达及TLR4、NF-κB p65蛋白阳性表达率显著降低。结论:虎杖苷可改善肺组织、支气管组织形态,降低大鼠气道炎症反应,其机制可能与抑制TLR4/NF-κB信号通路有关。
Objective: To explore the effect of polydatin on the airway inflammation in rats with chronic obstructive pulmonary disease(COPD) and its mechanism. Methods: Rats were randomly divided into the control group, COPD model group(model group), polydatin groups at doses of 30、60 and 120 mg/kg, 0.09 mg/kg dexamethasone group. Forced vital capacity(FVC), the forced expiratory volume in 0.1 second(FEV0.1), peak expiratory flow(PEF), maximum mid-expiratory flow(MMF) and complianc Lung dynamics(Cldyn) were measured with a pulmonary function detector. The total number of leukocytes and the proportion of polymorphonuclearleukocyte(PMN), alveolar macrophage(AM) and lymphocyte(LYM) cells in bronchoalveolar lavage fluid(BALF) were detected by Jimsa staining method. Levels of TNF-α, IL-8 and IL-1β in BALF were detected by ELISA. Pathological changes of lung tissues were observed by HE staining. Bronchial pathological changes were observed by Periodate Schiff's staining(PAS). Protein expressions of TLR4, NF-κB, matrix metalloproteinase-2(MMP-2) and MMP-9 in lung tissues were detected by Western blotting. Positive expression rates of TLR4 and NF-κB protein in lung tissues were detected by immunohistochemistry. Results: Compared with the control group, the pathological semi-quantitative scores of lung tissues in the model group were significantly increased. FVC, FEV0.1, PEF, MMF, Cldyn in the model group were decreased, AM cell proportion in BALF was decreased, the total number of white blood cell, PMN and LYM cell proportion was increased, levels of TNF-α, IL-8, IL-1βwere increased. The thickness of bronchial tracheal wall and smooth muscle were increased,protein expressions of TLR4, NF-κB p65, MMP-2, MMP-9 and protein positive expression rates of TLR4, NF-κB p65 in lung tissues were significantly increased. Compared with the model group,the pathological semi-quantitative scores of lung tissues in polydatin group and dexamethasone group were significantly decreased. FVC, FEV0.1, PEF, MMF and Cldyn in polydatin group and DXM group were increased, AM cell proportion in BALF was increased, the total number of white blood cell, PMN and LYM cell proportion was decreased,levels of TNF-α, IL-8, IL-1β were decreased. The thickness of bronchial tracheal wall and smooth muscle was decreased,protein expressions of TLR4, NF-κB p65, MMP-2, MMP-9 and protein expression rates of TLR4, NF-κB p65 in lung tissues were significantly decreased. Conclusion: Polydatin can improve the morphology of lung and bronchial tissues and reduce the airway inflammation in rats. Its mechanism may be related to the inhibition of TLR4/NF-kappa B signaling pathway.
Objective: To explore the effect of polydatin on the airway inflammation in rats with chronic obstructive pulmonary disease(COPD) and its mechanism. Methods: Rats were randomly divided into the control group, COPD model group(model group), polydatin groups at doses of 30、60 and 120 mg/kg, 0.09 mg/kg dexamethasone group. Forced vital capacity(FVC), the forced expiratory volume in 0.1 second(FEV0.1), peak expiratory flow(PEF), maximum mid-expiratory flow(MMF) and complianc Lung dynamics(Cldyn) were measured with a pulmonary function detector. The total number of leukocytes and the proportion of polymorphonuclearleukocyte(PMN), alveolar macrophage(AM) and lymphocyte(LYM) cells in bronchoalveolar lavage fluid(BALF) were detected by Jimsa staining method. Levels of TNF-α, IL-8 and IL-1β in BALF were detected by ELISA. Pathological changes of lung tissues were observed by HE staining. Bronchial pathological changes were observed by Periodate Schiff's staining(PAS). Protein expressions of TLR4, NF-κB, matrix metalloproteinase-2(MMP-2) and MMP-9 in lung tissues were detected by Western blotting. Positive expression rates of TLR4 and NF-κB protein in lung tissues were detected by immunohistochemistry. Results: Compared with the control group, the pathological semi-quantitative scores of lung tissues in the model group were significantly increased. FVC, FEV0.1, PEF, MMF, Cldyn in the model group were decreased, AM cell proportion in BALF was decreased, the total number of white blood cell, PMN and LYM cell proportion was increased, levels of TNF-α, IL-8, IL-1βwere increased. The thickness of bronchial tracheal wall and smooth muscle were increased,protein expressions of TLR4, NF-κB p65, MMP-2, MMP-9 and protein positive expression rates of TLR4, NF-κB p65 in lung tissues were significantly increased. Compared with the model group,the pathological semi-quantitative scores of lung tissues in polydatin group and dexamethasone group were significantly decreased. FVC, FEV0.1, PEF, MMF and Cldyn in polydatin group and DXM group were increased, AM cell proportion in BALF was increased, the total number of white blood cell, PMN and LYM cell proportion was decreased,levels of TNF-α, IL-8, IL-1β were decreased. The thickness of bronchial tracheal wall and smooth muscle was decreased,protein expressions of TLR4, NF-κB p65, MMP-2, MMP-9 and protein expression rates of TLR4, NF-κB p65 in lung tissues were significantly decreased. Conclusion: Polydatin can improve the morphology of lung and bronchial tissues and reduce the airway inflammation in rats. Its mechanism may be related to the inhibition of TLR4/NF-kappa B signaling pathway.
引文
[1]张弘,蔡柏蔷.慢性阻塞性肺疾病急性加重临床研究进展 .中国呼吸与危重监护杂志,2016,15(2)∶198～202.
[2]廖程程,陈亚红,林帆,等.皮质抑素在慢性阻塞性肺疾病大鼠中的变化及其意义 .中华医学杂志,2015,95(22)∶1722～1725.
[3]柳国洪,于洪江.他汀类药物对慢性阻塞性肺疾病大鼠气道炎症及黏液高分泌的作用及机制 .中国老年学杂志,2017,37(9)∶156～159.
[4]Wang X,Luo B,Lu Y,et al.The triggering receptor expressed by myeloid cells-1 activates TLR4-MyD88-NF-κB-dependent signaling to aggravate ventilation-induced lung inflammation and injury in mice .Cell Tissue Res,2018,374(1)∶137～148.
[5]封继宏,祁海燕,李美凤,等.扶正祛邪方对慢性阻塞性肺疾病稳定期模型大鼠肺组织细菌菌落的影响 .中医杂志,2016,57(4)∶337～341.
[6]Tang K S,Tan J S.The protective mechanisms of polydatin in cerebral ischemia.Eur J Pharmacol,2019,842(3)∶133～138.
[7]吴占庆,马强,冶国栋.虎杖苷对重症失血性休克患者心功能及外周血血小板线粒体的影响 .中国生化药物杂志,2015,35(9)∶146～148.
[8]黄杨,张新彧,孙纪元,等.虎杖苷对百草枯中毒大鼠急性肺损伤的保护作用 .临床急诊杂志,2017,18(3)∶182～184.
[9]张新彧,黄杨,孙纪元,等.虎杖苷对急性百草枯中毒大鼠肺损伤的干预研究 .解放军医药杂志,2018,30(4)∶246～249.
[10]宣肺平喘胶囊对慢性阻塞性肺疾病大鼠模型的影响 .中国实验方剂学杂志,2017,23(2)∶120～125.
[11]汪珊珊,汪电雷,陶秀华,等.脂多糖诱导的慢性阻塞性肺病模型大鼠肺支气管上皮MRP1功能分析 .中国实验动物学报,2014,22(3)∶30～34.
[12]Mikawa K,Nishina K,Takao Y,et al.ONO-1714,a nitric oxide synthase inhibitor,attenuates endotoxin-induced acute lung injury in rabbits .Anesth Analg,2003,97∶1751～1755
[13]张倩,黄萍,李艳,等.腹腔注射烟草烟雾提取物制备小鼠慢性阻塞性肺疾病模型的评价 .中华结核和呼吸杂志,2015,38(4)∶279～285.
[14]Zeng H,Wang Y,Gu Y.Polydatin attenuates reactive oxygen species-induced airway remodeling by promoting Nrf2-mediated antioxidant signaling in asthma mouse model.Life Sci,2018,27(8)∶598～607.
[15]Cao K,Lei X ,Liu H ,et al.Polydatin alleviated radiation-induced lung injury through activation of Sirt3 and inhibition of epithelial-mesenchymal transition .J Cell Mol Med,2017,21(12)∶3264～3276.
[16]王慧敏,戈改真,贺丽琼,等.幽门螺杆菌感染对慢性阻塞性肺疾病大鼠气道炎症因子的影响 .中国医药科学,2017,7(21)∶31～33.
[17]洪辉华,杨珺超,蔡宛如.芪冬活血饮对急性肺损伤大鼠Cav-1/NF-κB炎性反应信号通路的影响 .中华中医药杂志,2016,31(1)∶239～243.
[18]Tripathi P M ,Kant S ,Yadav R S ,et al.Expression of Toll-like Receptor 2 and 4 in Peripheral Blood Neutrophil Cells from Patients with Chronic Obstructive Pulmonary Disease.Oman Med J,2017,32(6)∶477～485.
[19]王鹏雁,蒋明,王昌明,等.脂多糖诱导下慢性阻塞性肺疾病大鼠模型远端肺动脉平滑肌细胞中Toll样受体4表达情况研究 .中国全科医学,2017,20(21)∶2603～2608.
[20]Huang B ,Liu J,Li Y,et al.Polydatin Prevents Lipopolysaccharide (LPS)-Induced Parkinson's Disease Via Regulation of the AKT/GSK3β-Nrf2/NF-ΚB Signaling Axis .Front Immunol,2018,9(11)∶2527～2536.
[21]Jiang Q ,Yi M ,Guo Q ,et al.Protective effects of polydatin on lipopolysaccharide-induced acute lung injury through TLR4-MyD88-NF-κB pathway .International Immunopharmacology,2015,29(2)∶370～376.
[22]Tang J,Li Y,Wang J ,et al.Polydatin suppresses the development of lung inflammation and fibrosis by inhibiting activation of the NACHT domain-,leucine-rich repeat-,and pyd-containing protein 3 inflammasome and the nuclear factor-κB pathway after Mycoplasma pneumoniae infection .J Cell Biochem,2018,67(10)∶6759～6768.
[23]Xie S S,Hu F,Tan M,et al.Relationship between expression of matrix metalloproteinase-9 and adenylyl cyclase-associated protein l in chronic obstructive pulmonary disease .J Int Med Res,2014,42(6)∶ 1272～1284
[24]Somaieh M ,Ali N ,Hassan G ,et al.The effect of conjugated linoleic acid on oxidative stress and matrix metalloproteinases 2 and 9 in patients with COPD .Int J Chron Obstruct Pulmon Dis,2018,13(5)∶1449～1454.
[2]廖程程,陈亚红,林帆,等.皮质抑素在慢性阻塞性肺疾病大鼠中的变化及其意义 .中华医学杂志,2015,95(22)∶1722～1725.
[3]柳国洪,于洪江.他汀类药物对慢性阻塞性肺疾病大鼠气道炎症及黏液高分泌的作用及机制 .中国老年学杂志,2017,37(9)∶156～159.
[4]Wang X,Luo B,Lu Y,et al.The triggering receptor expressed by myeloid cells-1 activates TLR4-MyD88-NF-κB-dependent signaling to aggravate ventilation-induced lung inflammation and injury in mice .Cell Tissue Res,2018,374(1)∶137～148.
[5]封继宏,祁海燕,李美凤,等.扶正祛邪方对慢性阻塞性肺疾病稳定期模型大鼠肺组织细菌菌落的影响 .中医杂志,2016,57(4)∶337～341.
[6]Tang K S,Tan J S.The protective mechanisms of polydatin in cerebral ischemia.Eur J Pharmacol,2019,842(3)∶133～138.
[7]吴占庆,马强,冶国栋.虎杖苷对重症失血性休克患者心功能及外周血血小板线粒体的影响 .中国生化药物杂志,2015,35(9)∶146～148.
[8]黄杨,张新彧,孙纪元,等.虎杖苷对百草枯中毒大鼠急性肺损伤的保护作用 .临床急诊杂志,2017,18(3)∶182～184.
[9]张新彧,黄杨,孙纪元,等.虎杖苷对急性百草枯中毒大鼠肺损伤的干预研究 .解放军医药杂志,2018,30(4)∶246～249.
[10]宣肺平喘胶囊对慢性阻塞性肺疾病大鼠模型的影响 .中国实验方剂学杂志,2017,23(2)∶120～125.
[11]汪珊珊,汪电雷,陶秀华,等.脂多糖诱导的慢性阻塞性肺病模型大鼠肺支气管上皮MRP1功能分析 .中国实验动物学报,2014,22(3)∶30～34.
[12]Mikawa K,Nishina K,Takao Y,et al.ONO-1714,a nitric oxide synthase inhibitor,attenuates endotoxin-induced acute lung injury in rabbits .Anesth Analg,2003,97∶1751～1755
[13]张倩,黄萍,李艳,等.腹腔注射烟草烟雾提取物制备小鼠慢性阻塞性肺疾病模型的评价 .中华结核和呼吸杂志,2015,38(4)∶279～285.
[14]Zeng H,Wang Y,Gu Y.Polydatin attenuates reactive oxygen species-induced airway remodeling by promoting Nrf2-mediated antioxidant signaling in asthma mouse model.Life Sci,2018,27(8)∶598～607.
[15]Cao K,Lei X ,Liu H ,et al.Polydatin alleviated radiation-induced lung injury through activation of Sirt3 and inhibition of epithelial-mesenchymal transition .J Cell Mol Med,2017,21(12)∶3264～3276.
[16]王慧敏,戈改真,贺丽琼,等.幽门螺杆菌感染对慢性阻塞性肺疾病大鼠气道炎症因子的影响 .中国医药科学,2017,7(21)∶31～33.
[17]洪辉华,杨珺超,蔡宛如.芪冬活血饮对急性肺损伤大鼠Cav-1/NF-κB炎性反应信号通路的影响 .中华中医药杂志,2016,31(1)∶239～243.
[18]Tripathi P M ,Kant S ,Yadav R S ,et al.Expression of Toll-like Receptor 2 and 4 in Peripheral Blood Neutrophil Cells from Patients with Chronic Obstructive Pulmonary Disease.Oman Med J,2017,32(6)∶477～485.
[19]王鹏雁,蒋明,王昌明,等.脂多糖诱导下慢性阻塞性肺疾病大鼠模型远端肺动脉平滑肌细胞中Toll样受体4表达情况研究 .中国全科医学,2017,20(21)∶2603～2608.
[20]Huang B ,Liu J,Li Y,et al.Polydatin Prevents Lipopolysaccharide (LPS)-Induced Parkinson's Disease Via Regulation of the AKT/GSK3β-Nrf2/NF-ΚB Signaling Axis .Front Immunol,2018,9(11)∶2527～2536.
[21]Jiang Q ,Yi M ,Guo Q ,et al.Protective effects of polydatin on lipopolysaccharide-induced acute lung injury through TLR4-MyD88-NF-κB pathway .International Immunopharmacology,2015,29(2)∶370～376.
[22]Tang J,Li Y,Wang J ,et al.Polydatin suppresses the development of lung inflammation and fibrosis by inhibiting activation of the NACHT domain-,leucine-rich repeat-,and pyd-containing protein 3 inflammasome and the nuclear factor-κB pathway after Mycoplasma pneumoniae infection .J Cell Biochem,2018,67(10)∶6759～6768.
[23]Xie S S,Hu F,Tan M,et al.Relationship between expression of matrix metalloproteinase-9 and adenylyl cyclase-associated protein l in chronic obstructive pulmonary disease .J Int Med Res,2014,42(6)∶ 1272～1284
[24]Somaieh M ,Ali N ,Hassan G ,et al.The effect of conjugated linoleic acid on oxidative stress and matrix metalloproteinases 2 and 9 in patients with COPD .Int J Chron Obstruct Pulmon Dis,2018,13(5)∶1449～1454.