摘要
目的 探讨连翘苷对呼吸道合胞病毒(RSV)感染小鼠Toll样受体4(TLR4)/髓样分化因子88(MyD88)信号通路的影响.方法 用RSV滴鼻感染法构建呼吸道感染模型,将RSV感染小鼠随机分为模型组、实验组及阳性对照组,另对照组小鼠给予等量的DMEM培养基滴鼻.实验组小鼠灌胃给予200 mg·kg-1·d-1连翘苷,阳性对照组小鼠灌胃给予27.6 mg·kg-1·d-1利巴韦林,对照组与模型组均灌胃给予等量0.9％NaCl,连续干预14 d.用酶联免疫吸附试验法检测肺泡灌洗液(BALF)中白细胞介素-4(IL-4)、IL-13及γ-干扰素(INF-γ)水平,用实时荧光定量聚合酶链反应法检测肺组织TLR4、MyD88 mRNA的转录水平,用免疫组织化学法检测肺组织TLR4、MyD88蛋白的阳性表达情况.结果 对照组、模型组、实验组及阳性对照组小鼠BALF中IL-4水平分别为(490.63±27.45)、(1 382.37±41.28)、(970.32±36.01)和(738.41±30.27)pg·mL-1,IL-13 水平分别为(4.02±2.63)、(18.33±5.62)、(12.97±3.73)和(7.51±2.74)pg·mL-1,INF-γ 水平分别为(54.42±2.68)、(30.26±4.68)、(42.80±3.36)和(45.01±3.82)pg·mL-1,肺组织 TLR4 mRNA 相对表达水平分别为 0.43±0.06、1.63±0.12、0.98±0.10 和 0.86±0.06,肺组织 MyD88 mRNA 相对表达水平分别为 0.56±0.09、1.78±0.15、1.02±0.07 和0.75±0.06,肺组织TLR4蛋白平均光密度值分别为0.02±0.00、0.11±0.02、0.06±0.01和0.03±0.00,MyD88蛋白平均光密度值分别为0.01±0.00、0.09±0.02、0.05±0.01和0.03±0.01.模型组的上述指标与对照组比较,实验组和阳性对照组的上述指标与模型组比较,在统计学上差异均有统计学意义(均P＜0.05).结论 连翘苷可减轻RSV感染小鼠的肺损伤,可能与调节免疫细胞的平衡,抑制TLR4/MyD88信号通路有关.
Abstract
Objective To explore the effect of forsythin on the Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)signaling pathway in mice infected with respiratory syncytial virus(RSV).Methods A respiratory infection model was established by intranasal inoculation of RSV.Mice infected with RSV were randomly divided into model group,experimental group,and positive control group,while control group received an equal volume of DMEM medium by nasal drops.The experimental group was administered forsythin(200 mg·kg-1·d-1)by gavage,the positive control group received ribavirin(27.6 mg·kg-1·d-1)by gavage,and both the control and model groups were given an equal volume of 0.9％NaCl,with continuous intervention for 14 days.Interleukin-4(IL-4),IL-13,and interferon-gamma(INF-γ)levels in bronchoalveolar lavage fluid(BALF)were detected by enzyme-linked immunosorbent assay;transcription levels of TLR4 and MyD88 mRNA in lung tissue were detected by real-time fluorescent quantitative polymerase chain reaction;and the positive expression of TLR4 and MyD88 proteins in lung tissue were detected by immunohistochemistry.Results The levels of IL-4 in BALF of the control group,model group,experimental group,and positive control group were(490.63±27.45),(1 382.37±41.28),(970.32±36.01),(738.41±30.27)pg·mL-1,respectively;IL-13 levels were(4.02±2.63),(18.33±5.62),(12.97±3.73),and(7.51±2.74)pg·mL-1,respectively;INF-γ levels were(54.42±2.68),(30.26±4.68),(42.80±3.36),and(45.01±3.82)pg·mL-1,respectively;the expression of TLR4 mRNA in lung tissue were 0.43±0.06,1.63±0.12,0.98±0.10,and 0.86±0.06,respectively;the expression of MyD88 mRNA in lung tissue were 0.56±0.09,1.78±0.15,1.02±0.07,and 0.75±0.06,respectively;the average optical density of TLR4 protein in lung tissue were 0.02±0.00,0.11±0.02,0.06±0.01,and 0.03±0.00,respectively;the average optical density of MyD88 protein were respectively 0.01±0.00,0.09±0.02,0.05±0.01,and 0.03±0.01,respectively.Compared the model group with the control group,and compared the experimental and positive control groups with the model group,above indicators showed statistically significant differences(all P＜0.05).Conclusion Forsythin can alleviate lung damage in mice infected with RSV,possibly by regulating the balance of immune cells and inhibiting the TLR4/MyD88 signaling pathway.
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