Effects of Qingfei Granule protecting airway injury of COPD rats by TLR4/NF⁃κB signaling pathway

Zhe ZHE; Jianshan LIAO; Wenjie LI; Jiayu LIU; Zhixiang ZHAO; Dan LIU; Binqing TANG; Yada ZHANG; Bidan XIONG; Kehua SHI

doi:10.16305/j.1007-1334.2022.2007024

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Effects of Qingfei Granule protecting airway injury of COPD rats by TLR4/NF⁃κB signaling pathway

更新时间：2022-07-10

- Effects of Qingfei Granule protecting airway injury of COPD rats by TLR4/NF⁃κB signaling pathway
- Shanghai Journal of Traditional Chinese Medicine Vol. 56, Issue 3, Pages: 64-73(2022)
- 作者机构：
  
  1.上海中医药大学附属市中医医院（上海 200071）
  2.上海中医药大学（上海 201203）
  3.静安区北站医院（上海　200070）
- 作者简介：
- 基金信息：
- DOI：10.16305/j.1007-1334.2022.2007024
  CLC：
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折哲,廖健杉,李文杰等.清肺颗粒干预TLR4/NF⁃κB信号通路调控COPD大鼠气道损伤的机制研究[J].上海中医药杂志,2022,56(03):64-73.

ZHE Zhe,LIAO Jianshan,LI Wenjie,et al.Effects of Qingfei Granule protecting airway injury of COPD rats by TLR4/NF⁃κB signaling pathway[J].Shanghai Journal of Traditional Chinese Medicine,2022,56(03):64-73.
折哲,廖健杉,李文杰等.清肺颗粒干预TLR4/NF⁃κB信号通路调控COPD大鼠气道损伤的机制研究[J].上海中医药杂志,2022,56(03):64-73. DOI： 10.16305/j.1007-1334.2022.2007024.

ZHE Zhe,LIAO Jianshan,LI Wenjie,et al.Effects of Qingfei Granule protecting airway injury of COPD rats by TLR4/NF⁃κB signaling pathway[J].Shanghai Journal of Traditional Chinese Medicine,2022,56(03):64-73. DOI： 10.16305/j.1007-1334.2022.2007024.

摘要

目的,2,从气道炎症和气道屏障损伤角度探索清肺颗粒通过干预TLR4/NF-κB信号通路调控慢性阻塞性肺疾病（COPD）大鼠气道损伤的作用机制。,方法,2,SD大鼠60只，随机选取10只为正常组，其余50只大鼠均采用香烟烟雾（CS）暴露联合气管注射脂多糖（LPS）的方法建立COPD模型。造模成功后，将上述50只大鼠随机分为模型组，地塞米松组（灌胃地塞米松0.2 mg/kg），清肺低、中、高剂组（分别灌胃清肺颗粒5.4 g/kg、10.8 g/kg、21.4 g/kg），每组10只，共灌胃8周。测定大鼠肺功能变化情况，透射电镜下观察肺脏亚显微结构改变；ELISA法测定血清及支气管肺泡灌洗液（BALF）中细胞因子［包括转化生长因子β1（TGF-β1）、肿瘤坏死因子-α（TNF-α）、白介素-1β（IL-1β）、白介素-6（IL-6）、白介素-10（IL-10）、血管内皮生长因子（VEGF）、结缔组织生长因子（CTGF）、内皮素-1（ET-1）］含量；qRT-PCR法测定肺组织中微小RNA-146a（,miRNA-146a,）、Toll样受体4（,TLR4）、,核转录因子⁃κB（,NF⁃κB,）,、,B淋巴细胞瘤⁃2基因（,Bcl⁃2,）、Bcl⁃2相关X蛋白,（Bax,）mRNA的表达情况；Western blot法测定肺组织TLR4、NF-κB、Bcl-2、Bax的蛋白表达水平。,结果,2,①与模型组比较，清肺颗粒各组的肺通气功能有所改善，表现为0.2 s用力呼气容积（FEV0.2）/用力肺活量（FVC）水平均有所上升（,P<,0.05），而RL和Re水平有不同程度下降（,P<,0.05），且清肺高剂组对大鼠肺功能FEV0.2和FEV0.2/FVC水平的改善程度与地塞米松组相似（,P>,0.05）。②通过透射电子显微镜下观察，与模型组比较，清肺高剂组的Ⅱ型肺泡上皮细胞胞浆质地均匀，核膜未见明显破坏，可见较多线粒体和板层小体，细胞形态大多正常。③与模型组比较，清肺颗粒可降低大鼠血清和BALF液中TNF-α、IL-8水平（,P<,0.05），且中、高剂量组具有一定的剂量依赖关系。④与模型组比较，清肺颗粒可上调,miRNA,-,146a, mRNA表达，降低大鼠肺组织,TLR4、NF,-,κB、Bax, mRNA和其蛋白表达量，上调,Bcl,-,2, mRNA和其蛋白表达量（,P<,0.05）。,结论,2,清肺颗粒可通过抑制TLR4/NF-κB信号通路相关mRNA及蛋白表达，抑制肺泡上皮细胞凋亡，进而达到修复气道屏障损伤的作用。

Abstract

Objective,2,To explore the mechanism of Qingfei Granule regulating airway injury in COPD rats by interfering with TLR4/NF- κB signal pathway from the point of view of airway inflammation and airway barrier injury.,Methods,2,In 60 SD rats， 10 rats were randomly selected as the normal group， and the other 50 rats were exposed to cigarette smoke combined with endotracheal injection of LPS to establish COPD model. After successful establishment of the model， 50 rats were randomly divided into the model group， dexamethasone group （dexamethasone 0.2 mg/kg）， Qingfei Granule low， medium and high dose groups （Qingfei Granule 5.4 g/kg， 10.8 g/kg， 21.4 g/kg， respectively）， with 10 rats in each group and were respectively given the corresponding drugs by intragastric administration for 8 weeks. The changes of pulmonary function and ultrastructure of lung were observed under transmission electron microscope. The contents of cytokines （including TGF- β1， TNF- α， IL-1β， IL-6， IL-10， VEGF， CTGF and ET-1） in serum and bronchoalveolar lavage fluid （BALF） were measured by ELISA. The expressions of ,TLR4,， ,NF⁃κB,， ,Bcl⁃2, and ,Bax, mRNA in lung tissue were measured by qRT-PCR. The protein expressions of TLR4， NF-κB， Bcl-2 and Bax in lung tissue were measured by Western blot.,Results,2,①Compared with the model group， the pulmonary ventilation function of different Qingfei Granule groups was improved， showing that the level of FEV0.2/FVC was increased （,P,<,0.05）， while the levels of RL and Re decreased in different degrees. The improvement degree of FEV0.2 and FEV0.2/FVC in high dose group of Qingfei Granule was similar to that in dexamethasone group（,P,>,0.05）.②Through the observation under transmission electron microscope， compared with the model group， the cytoplasmic texture of type Ⅱ alveolar epithelial cells in the high dose group of Qingfei Granule was uniform. There was no obvious destruction of nuclear membrane. More mitochondria and lamellar bodies could be seen， and most of the cells were normal. ③Compared with the model group， Qingfei Granule could reduce the levels of TNF-α and IL-8 in serum and BALF fluid of rats（,P<,0.05）， and the medium and the high dose groups had a certain dose-dependent relationship. ④Compared with the model group， Qingfei Granule could increased the expression of ,miRNA⁃146a, mRNA， reduce the expression of ,TLR4,， ,NF⁃κB,， ,Bax, mRNA and its protein， increased the expression of ,Bcl⁃2 ,mRNA and its protein in lung tissue of rats（,P<,0.05）.,Conclusion,2,Qingfei Granule can repair the airway barrier injury by inhibiting the expression of mRNA and protein related to TLR4/NF-κB signal pathway and inhibiting the apoptosis of alveolar epithelial cells.

关键词

TLR4/NF-κB信号通路凋亡与修复机制研究模型大鼠中药研究

Keywords

TLR4/NF-κB signaling pathwayapoptosis and repairmechanism studymodel ratstraditional Chinese herbal medicine research

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