苍耳子挥发油对支气管哮喘大鼠气道重塑的影响
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摘要
目的:探讨苍耳子挥发油对支气管哮喘模型大鼠气道重塑的影响,并从对基质金属蛋白酶-9(MMP-9),基质金属蛋白酶组织抑制剂-1(TIMP-1),血小板衍生生长因子(PDGF),转化生长因子-β_1(TGF-β_1),Smad2,Smad3和Smad7 mRNA的表达方面探讨了其作用机制。方法:SD大鼠雌雄各半,共48只,随机分为正常组,模型组,苍耳子挥发油7. 5,15,30 mg·kg-1组和地塞米松(0. 5 mg·kg~(-1))组,每组各8只。除正常组外,用卵蛋白致敏并雾化吸入法诱发大鼠支气管哮喘模型。苍耳子挥发油给药组从第1次哮喘激发开始(造模后第4周)至处死前每天通过喷雾给予苍耳子挥发油干预,共4周。进行苏木素-伊红(HE)染色分析支气管肺组织病理学改变;进行气道重塑情况观察,记录支气管管腔内周长(Pi),支气管平滑肌面积(S),管壁面积(W),平滑肌细胞核数(N);检测肺泡灌洗液(BALF)中内皮素-1(ET-1)和类胰岛素生长因子-1(IGF-1)水平;采用实时荧光定量聚合酶链式反应(Real-time PCR)法检测肺组织MMP-9,TIMP-1,PDGF,TGF-β_1,Smad2,Smad3和Smad7 mRNA的表达。结果:模型组支气管壁周围有大量炎性细胞浸润,气道平滑肌厚度明显增加,官腔变窄扁平,基底膜增厚;苍耳子挥发油各剂量组明显改善支气管肺组织病理损伤。与正常组比较,模型组S/Pi,W/Pi,N/Pi均明显升高(P <0. 01),苍耳子挥发油各剂量组和地塞米松组S/Pi,W/Pi,N/Pi均低于模型组(P <0. 01)。与正常比较,模型组BALF中ET-1和IGF-1水平明显升高(P <0. 01),苍耳子挥发油各剂量组和地塞米松组BALF中ET-1和IGF-1水平明显低于模型组(P <0. 01)。与正常比较,模型组MMP-9 mRNA表达明显升高(P <0. 01),TIMP-1 mRNA表达明显降低(P <0. 01),MMP-9/TIMP-1明显升高(P <0. 01),苍耳子挥发油各剂量组和地塞米松组MMP-9 mRNA表达和MMP-9/TIMP-1低于模型组(P <0. 01)。与正常组比较,模型组肺组织PDGF mRNA表达明显增强(P <0. 01),苍耳子挥发油各剂量组和地塞米松组PDGF mRNA表达均低于模型组(P <0. 01)。与正常组相比较,模型组TGF-β1,Smad2和Smad3 mRNA表达均增强(P <0. 01),Smad7 mRNA表达减弱(P <0. 01),与模型组比较,苍耳子挥发油各剂量组和地塞米松组TGF-β_1,Smad2和Smad3 mRNA表达减弱(P <0. 01),Smad7 mRNA表达增强(P <0. 01)。结论:苍耳子挥发油可调节MMP-9,TIMP-1水平及MMP-9/TIMP-1值,调节TGF-β1/Smad信号通路,抑制PDGF,ET-1和IGF-1因子表达,通过多种途径抑制了上皮下纤维化、细胞外基质合成、气道平滑肌细胞增殖、迁移等,起到减轻或抑制气道重塑,从而起到防治哮喘的作用。
Objective: To discuss the effect of volatile oil from Xanthii Fructus on airway remodeling of rats with bronchial asthma,in order to discuss the mechanism of action through matrix metalloteinases-9( MMP-9),tissue inhibitor of metalloproteinase-1( TIMP-1),platelet derived growth factor( PDGF),transforming growth factor-β_1( TGF-β_1),Smad2,Smad3 and Smad7 mRNA expressions. Method: Forty-eight rats( half males and half females) were randomly divided into six groups( n = 8),namely normal saline group,model group,volatile oil from Xanthii Fructus( 7. 5,15,30 mg·kg~(-1)) groups and dexamethasone group( 0. 5 mg·kg~(-1)). Except for normal saline group,airway remodeling of rats were established through ovalbumin sensitization and atomization inhalation method. At volatile oil group,rats got volatile oil spray since the first time( the 4 thweek after modeling)of inspiration of asthma to the last day before the end of test. Histopathological changes of bronchus and lung were analyzed by htoxylin eosin( HE) stain. And airway remodeling was observed,and perimeter of bronchial lumen( Pi),smooth muscle area of bronchi( S),tube wall area( W) and nuclei of smooth muscle cells( N) were recorded. And levels of endothelin-1( ET-1) and insulin-like growth factor-1( IGF-1) in irrigation solution of Alveoli( BALF) were detected. And mRNA expressions of MMP-9,TIMP-1,PDGF,TGF-β1,Smad2,Smad3 and Smad7 were detected by real-time quantitative-polymerase chain reaction( Real-time PCR). Result: In the model group,infiltration of a large number of inflammatory cells were found at the peribronchial wall,airway smooth muscle thickness and basement membrane increased,and bureaucratic cavity was narrower and flatter. Volatile oil from Xanthii Fructus can ameliorate pathological damage of bronchopulmonary tissue. Levels of S/Pi,W/Pi and N/Pi,BALF,ET-1,IGF-1 and MMP-9/TIMP-1 and mRNA expressions of MMP-9 and PDGF in volatile oil group and dexamethasone group were lower than those in model group( P < 0. 01). Compared with model group,mRNA expressions of TGF-β1,Smad2 and Smad3 were less than those in model group( P < 0. 01),while mRNA expression of Smad7 was more than that in model group( P < 0. 01). Conclusion: Volatile oil from Xanthii Fructus can regulate levels of MMP-9,TIMP-1,MMP-9/TIMP-1,TGF-β1/Smad,inhibit expressions of PDGF,ET-1 and IGF-1,and suppress subepithelial fibrosis,extracellular matrix synthesis,proliferation and migration of airway smooth muscle cells to relieve or inhibit airway remodeling,so as to prevent and treat asthma.
Objective: To discuss the effect of volatile oil from Xanthii Fructus on airway remodeling of rats with bronchial asthma,in order to discuss the mechanism of action through matrix metalloteinases-9( MMP-9),tissue inhibitor of metalloproteinase-1( TIMP-1),platelet derived growth factor( PDGF),transforming growth factor-β_1( TGF-β_1),Smad2,Smad3 and Smad7 mRNA expressions. Method: Forty-eight rats( half males and half females) were randomly divided into six groups( n = 8),namely normal saline group,model group,volatile oil from Xanthii Fructus( 7. 5,15,30 mg·kg~(-1)) groups and dexamethasone group( 0. 5 mg·kg~(-1)). Except for normal saline group,airway remodeling of rats were established through ovalbumin sensitization and atomization inhalation method. At volatile oil group,rats got volatile oil spray since the first time( the 4 thweek after modeling)of inspiration of asthma to the last day before the end of test. Histopathological changes of bronchus and lung were analyzed by htoxylin eosin( HE) stain. And airway remodeling was observed,and perimeter of bronchial lumen( Pi),smooth muscle area of bronchi( S),tube wall area( W) and nuclei of smooth muscle cells( N) were recorded. And levels of endothelin-1( ET-1) and insulin-like growth factor-1( IGF-1) in irrigation solution of Alveoli( BALF) were detected. And mRNA expressions of MMP-9,TIMP-1,PDGF,TGF-β1,Smad2,Smad3 and Smad7 were detected by real-time quantitative-polymerase chain reaction( Real-time PCR). Result: In the model group,infiltration of a large number of inflammatory cells were found at the peribronchial wall,airway smooth muscle thickness and basement membrane increased,and bureaucratic cavity was narrower and flatter. Volatile oil from Xanthii Fructus can ameliorate pathological damage of bronchopulmonary tissue. Levels of S/Pi,W/Pi and N/Pi,BALF,ET-1,IGF-1 and MMP-9/TIMP-1 and mRNA expressions of MMP-9 and PDGF in volatile oil group and dexamethasone group were lower than those in model group( P < 0. 01). Compared with model group,mRNA expressions of TGF-β1,Smad2 and Smad3 were less than those in model group( P < 0. 01),while mRNA expression of Smad7 was more than that in model group( P < 0. 01). Conclusion: Volatile oil from Xanthii Fructus can regulate levels of MMP-9,TIMP-1,MMP-9/TIMP-1,TGF-β1/Smad,inhibit expressions of PDGF,ET-1 and IGF-1,and suppress subepithelial fibrosis,extracellular matrix synthesis,proliferation and migration of airway smooth muscle cells to relieve or inhibit airway remodeling,so as to prevent and treat asthma.
引文
[1]祝丁,张超,赖天文,等.中药对支气管哮喘气道重塑干预机制研究进展[J].中国实用内科杂志,2016,36(8):694-697.
[2]潘亦林,朱燕亭,李满祥.支气管哮喘气道重塑的研究进展[J].中华肺部疾病杂志:电子版,2015,8(6):773-776.
[3] Hirota N,Martin J G. Mechanisms of airway remodeling[J]. Chest,2013,144(3):1026-1032.
[4]崔秀荣,马海波,张旗,等.苍耳子的化学成分和临床应用研究进展[J].现代药物与临床,2012,27(6):614-618.
[5]李景福.辛夷、苍耳子对支气管哮喘患者Th1/Th2比值及炎性递质的影响[J].现代中西医结合杂志,2012,21(10):1057-1058.
[6]朱慧华,虞坚尔,陈燕萍,等.辛夷苍耳子治疗儿童哮喘缓解期慢性气道炎症机理研究[J].辽宁中医杂志,2007,34(8):1025-1028.
[7]黄钢花,黎凯燕,黎世明,等.小儿喘咳液合苍耳子散对哮喘大鼠气道重塑影响的研究[J].中华中医药学刊,2016,34(7):1682-1685.
[8]汝绍刚,赵文英,朱庆书,等.苍耳子油回流提取工艺优化及其动力学研究[J].化学工业与工程,2012,29(1):34-38.
[9]李翎,邹衍衍,石琛,等.β、α细辛醚及石菖蒲挥发油对支气管哮喘的药效对比观察[J].时珍国医国药,2006,17(11):2137-2138.
[10]张雄飞,黄娟萍,李碧云,等.中药治疗哮喘作用机制的研究进展[J].中国实验方剂学杂志,2013,19(15):344-347.
[11]乡世健,盛华芳,吕亚梅,等.冬病夏治方穴位贴敷对哮喘豚鼠气道炎症及肠道菌群的影响[J].中国实验方剂学杂志,2018,24(9):95-102.
[12]王忠敏,汤卫红,王惠庭,等.哮喘模型小鼠肺组织TGF-β1、MMP-9、TIMP-1与气道重塑相关性研究[J].浙江中西医结合杂志,2018,28(11):920-923.
[13]宋洪娟,黄正桥,黄笑,等.清金化痰汤通过p38MAPK/NF-κB信号通路改善大鼠急性气道炎症的作用和机制[J].中国实验方剂学杂志,2017,23(13):104-110.
[14]邹璐,孙祝美,郭春荣,等.中医药防治支气管哮喘气道重塑相关信号通路研究进展[J].中华中医药杂志,2018,33(11):5057-5060.
[15]杨季国,马慧娟,徐缨,等.平喘Ⅰ号对呼吸道合胞病毒哮喘鼠气道重塑PDGF-B与ERK信号通路的影响[J].中华中医药学刊,2014,32(11):2696-2699.
[16]吕昊泽,杨斌,康振华.血浆TNF-α与ET-1与肺炎支原体感染合并支气管哮喘相关性研究[J].中国实验诊断学,2016,20(9):1539-1540.
[17] Ciesielska N,Kozakiewicz M,Zukow W. Influence of physical activity on the levels IGF-1 and IGFBPs at geriatric patients. Review of clinical trials[J]. J Thorac Cardiovasc Surg,2015,150(2):358-366.
[18]王相东.苍耳子浸出液治疗226例变异性哮喘疗效观察[J].中国疗养医学,2016,25(3):312-313.
[19]张婷婷,鄢良春,赵军宁,等.苍耳子"毒性"及现代毒理学研究进展[J].药物评价研究,2010,33(5):361-366.
[2]潘亦林,朱燕亭,李满祥.支气管哮喘气道重塑的研究进展[J].中华肺部疾病杂志:电子版,2015,8(6):773-776.
[3] Hirota N,Martin J G. Mechanisms of airway remodeling[J]. Chest,2013,144(3):1026-1032.
[4]崔秀荣,马海波,张旗,等.苍耳子的化学成分和临床应用研究进展[J].现代药物与临床,2012,27(6):614-618.
[5]李景福.辛夷、苍耳子对支气管哮喘患者Th1/Th2比值及炎性递质的影响[J].现代中西医结合杂志,2012,21(10):1057-1058.
[6]朱慧华,虞坚尔,陈燕萍,等.辛夷苍耳子治疗儿童哮喘缓解期慢性气道炎症机理研究[J].辽宁中医杂志,2007,34(8):1025-1028.
[7]黄钢花,黎凯燕,黎世明,等.小儿喘咳液合苍耳子散对哮喘大鼠气道重塑影响的研究[J].中华中医药学刊,2016,34(7):1682-1685.
[8]汝绍刚,赵文英,朱庆书,等.苍耳子油回流提取工艺优化及其动力学研究[J].化学工业与工程,2012,29(1):34-38.
[9]李翎,邹衍衍,石琛,等.β、α细辛醚及石菖蒲挥发油对支气管哮喘的药效对比观察[J].时珍国医国药,2006,17(11):2137-2138.
[10]张雄飞,黄娟萍,李碧云,等.中药治疗哮喘作用机制的研究进展[J].中国实验方剂学杂志,2013,19(15):344-347.
[11]乡世健,盛华芳,吕亚梅,等.冬病夏治方穴位贴敷对哮喘豚鼠气道炎症及肠道菌群的影响[J].中国实验方剂学杂志,2018,24(9):95-102.
[12]王忠敏,汤卫红,王惠庭,等.哮喘模型小鼠肺组织TGF-β1、MMP-9、TIMP-1与气道重塑相关性研究[J].浙江中西医结合杂志,2018,28(11):920-923.
[13]宋洪娟,黄正桥,黄笑,等.清金化痰汤通过p38MAPK/NF-κB信号通路改善大鼠急性气道炎症的作用和机制[J].中国实验方剂学杂志,2017,23(13):104-110.
[14]邹璐,孙祝美,郭春荣,等.中医药防治支气管哮喘气道重塑相关信号通路研究进展[J].中华中医药杂志,2018,33(11):5057-5060.
[15]杨季国,马慧娟,徐缨,等.平喘Ⅰ号对呼吸道合胞病毒哮喘鼠气道重塑PDGF-B与ERK信号通路的影响[J].中华中医药学刊,2014,32(11):2696-2699.
[16]吕昊泽,杨斌,康振华.血浆TNF-α与ET-1与肺炎支原体感染合并支气管哮喘相关性研究[J].中国实验诊断学,2016,20(9):1539-1540.
[17] Ciesielska N,Kozakiewicz M,Zukow W. Influence of physical activity on the levels IGF-1 and IGFBPs at geriatric patients. Review of clinical trials[J]. J Thorac Cardiovasc Surg,2015,150(2):358-366.
[18]王相东.苍耳子浸出液治疗226例变异性哮喘疗效观察[J].中国疗养医学,2016,25(3):312-313.
[19]张婷婷,鄢良春,赵军宁,等.苍耳子"毒性"及现代毒理学研究进展[J].药物评价研究,2010,33(5):361-366.