Effect of Xiyanping on Inflammatory Microenvironment of COPD Model Based on TLR3-MAPK Signaling Pathway
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摘要:
目的 基于Toll样受体3蛋白（toll-like receptor 3，TLR3）-丝裂原活化蛋白激酶（mitogen-activated protein kinase，MAPK）信号通路探讨喜炎平（XYP）对慢性阻塞性肺病（chronic obstructive pulmonary disease，COPD）模型炎症微环境的影响。 方法 建立香烟烟雾（cigarette smoke，CS）诱导的COPD小鼠模型，采用XYP（低、中、高）进行治疗。将人支气管上皮细胞（human bronchial epithelial cells，HBE）暴露于香烟烟雾提取物（cigarette smoke extract，CSE），并用XYP或TLR3激动剂poly（I:C）进行处理。通过蛋白质印迹法检测α-平滑肌肌动蛋白（α-Smooth Muscle Actin，α-SMA）、I型胶原（collagen type I，CollagenⅠ）、TLR3、MAPK信号通路蛋白表达。 结果 与Ctrl组相比，CS组小鼠HE染色评分升高（P < 0.05）；与CS组相比，XYP-H组小鼠HE染色评分降低（P < 0.05）。与Ctrl组相比，CS组小鼠肺组织中CollagenⅠ、α-SMA、TLR3及MAPK蛋白水平升高（P < 0.05）；与CS组相比，XYP降低CollagenⅠ、α-SMA、TLR3及MAPK蛋白表达（P < 0.05）。体外实验中，与CSE组相比，CSE+XYP-H组HBE细胞中CollagenⅠ、α-SMA、TLR3蛋白表达及MAPK信号通路蛋白水平降低（P < 0.05）。与CSE+XYP-H组相比，CSE+XYP-H+poly（I:C）组中CollagenⅠ、α-SMA表达水平升高（P < 0.05）。 结论 喜炎平注射液减轻了COPD进程中肺纤维化的发展，其机制可能与抑制TLR3-MAPK信号通路的激活相关。 Abstract:Objective Based on Toll-like receptor 3 protein （TLR3）- mitogen-activated protein kinase （MAPK） signaling pathway, the effect of Xiyanping on inflammatory microenvironment of chronic obstructive pulmonary disease （COPD） model was discussed. Methods A mouse model of COPD induced by Cigarette Smoke （CS） was established and treated with XYP （low, medium and high）. Human Bronchial Epithelial cells （HBE） were exposed to Cigarette Smoke Extract （CSE） and treated with XYP or TLR3 agonist poly（I:C）. The expression of α-Smooth Muscle Actin （α-SMA）, Collagen type I （Collagen I）, TLR3 and MAPK signaling pathway proteins were detected by western blot. Results Compared with the Ctrl group, the score of HE staining in CS group increased （P < 0.05）. Compared with CS group, the score of HE staining in XYP-H group decreased （P < 0.05）. Compared with the Ctrl group, the levels of CollagenⅠ, α-SMA, TLR3 and MAPK signaling pathway protein in lung tissue of CS group increased （P < 0.05）. Compared with CS group, the expression of CollagenⅠ, α-SMA, TLR3 and MAPK protein in XYP-H group decreased （P < 0.05）. In vitro experiment, compared with CSE group, the expression of CollagenⅠ, α-SMA, TLR3 protein and MAPK signaling pathway protein in HBE cells in CSE+XYP-H group decreased （P < 0.05）. Compared with CSE+XYP-H group, the expression levels of CollagenⅠ and α-SMA in CSE+XYP-H+poly（I:C） group increased （P < 0.05）. Conclusion Xiyanping injection can alleviate the development of pulmonary fibrosis in the process of COPD, and its mechanism may be related to inhibiting the activation of TLR3-MAPK signaling pathway. -
图 1 小鼠肺组织的HE染色（$\bar x \pm s $，n = 6）
A：HE染色代表图；B：HE评分结果；标尺=50 μm；与CS组相比，**P < 0.01。
Figure 1. HE staining of mouse lung tissue（$\bar x \pm s $，n = 6）
图 2 血清学指标和肺功能指标（$\bar x \pm s $，n = 6）
A：小鼠CS模型肺功能参数；B：小鼠CS模型血清IL-6表达情况；C：小鼠CS模型血清TNF-α表达情况；与CS组相比，**P < 0.01。
Figure 2. Serological index and pulmonary function index（$\bar x \pm s $，n = 6）
图 3 小鼠肺组织CollagenⅠ、α-SMA蛋白表达（$\bar x \pm s $，n = 6）
A：CollagenⅠ、α-SMA蛋白的电泳条带图；B：CollagenI和α-SMA蛋白表达统计学分析；与CS组相比，**P < 0.01。
Figure 3. CollagenI and α-SMA protein expression in mouse lung tissue（$\bar x \pm s $，n = 6）
图 4 小鼠肺组织TLR3-MAPK信号通路蛋白表达（$\bar x \pm s $，n = 3）
A：TLR3-MAPK信号通路蛋白电泳条带图；B：TLR3-MAPK信号通路蛋白表达统计学分析；与CS组或XYP-M组相比，*P < 0.05，**P < 0.01。
Figure 4. TLR3-MAPK signaling pathway protein expression in mouse lung tissue（$\bar x \pm s $，n = 3）
图 5 HBE细胞纤维化指标分析（$\bar x \pm s $，n = 3）
A：CollagenⅠ、α-SMA蛋白的电泳条带图；B：CollagenI和α-SMA蛋白表达统计学分析；与CSE组或CSE+XYP-M组相比，*P < 0.05，**P < 0.01。
Figure 5. Analysis of HBE cell fibrosis indicators （$\bar x \pm s $，n = 3）
图 6 暴露于CES的HBE细胞中TLR3-MAPK信号通路蛋白表达（$\bar x \pm s $，n = 3）
A：TLR3-MAPK信号通路蛋白电泳条带图；B：TLR3-MAPK信号通路蛋白表达统计学分析；与CSE组或CSE+XYP-M组相比，*P < 0.05，**P < 0.01。
Figure 6. TLR3-MAPK signaling pathway protein expression in HBE cells exposed to CES（$\bar x \pm s $，n = 3）
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