清肺口服液对呼吸道合胞病毒感染小鼠内炎性细胞及Treg/Th17表达水平的影响

Effects of Qingfei Oral Liquid on the expressions of IL-10 and IL-17 in the lung tissue and those of Treg and Th17 in the spleen of RSV-infected mice

目的清肺口服液治疗呼吸道合胞病毒(respiratory syncytial viral,RSV)肺炎的作用机理研究尚不够深入。文中旨在通过检测清肺口服液治疗前后感染RSV肺炎小鼠肺组织内IL-10、IL-17的表达水平以及脾组织细胞中Treg、Th17细胞因子表达含量,从Treg/Th17细胞平衡角度对其作用机制进行探讨。方法将50只BABL/c小鼠随机分为5组,每组10只:对照组、模型组、利巴韦林组、清肺低剂量组、清肺高剂量组,除对照组予等量等渗盐水滴鼻外,其余4组用RSV(Long株)滴鼻造模,病毒激发48 h后,各组小鼠均灌胃给药,每日1次,对照组和模型组给予0.9%等渗盐水,清肺高剂量组、清肺低剂量组、利巴韦林组分别给予清肺口服液4 g/m L、清肺口服液1.33 g/m L、利巴韦林0.0025 g/m L,给药72 h后处死小鼠,每组取5只小鼠肺组织行病理分析,另外5只用ELISA方法检测肺泡灌洗液中IL-10、IL-17的含量,每组取4只用流式细胞术方法检测小鼠脾组织细胞Treg、Th17细胞因子表达含量。结果清肺高剂量组、清肺低剂量组、利巴韦林组、对照组病理改变评分较模型组降低(P<0.01);且IL-10表达[(46.64±0.48)、(43.54±1.03)、(47.83±0.87)、(50.44±1.04)ng/L]较模型组[(39.21±1.57)ng/L]均明显升高(P<0.01);而IL-17表达[(33.66±0.70)、(55.92±0.83)、(21.92±1.38)、(9.42±0.59)pg/m L]较模型组[(70.96±0.53)pg/m L]均明显降低(P<0.01)。清肺高、低剂量组IL-10、IL-17表达分别较利巴韦林组[(47.83±0.87)ng/L、(21.92±1.38)pg/m L]降低和升高(P<0.05)。清肺高剂量组、清肺低剂量组、利巴韦林组及对照组Treg/Th17相对表达[(6.38±0.36)、(2.89±0.52)、(3.95±0.26)、(3.54±0.85)]较模型组(0.96±0.16)均明显升高(P<0.01);且清肺高、低剂量组Treg、Th17、Treg/Th17表达与利巴韦林组比较的差异有统计学意义(P<0.05)。结论清肺口服液具有调节Treg/Th17平衡,并对于感染RSV小鼠体内IL-10细胞因子具有明显提升作用,而对IL-17细胞因子具有明显降低作用,可证明清肺口服液治疗RSV病毒性肺炎的有效性。

Objective The action mechanisms of Qingfei Oral Liquid（ QFOL） in the treatment of respiratory syncytial virus（ RSV） infection need to be studied more deeply. The aim of this study is to examine the expressions of interleukin（ IL）-10 and IL-17 in the lung tissue and those of Treg and Th17 in the spleen tissue of RSVinfected mice before and after treated with QFOL,and to explore the action mechanisms of QFOL from the perspective of the Treg/Th17 cytokines balance. Methods Fifty BABL/c mice were equally randomized to five groups： blank control,RSV model,Ribavirin,lowdose QFOL,and high-dose QFOL. Models of RSV（ long strain） infection were made in the latter four groups. At 48 hours after viral activation,the mice of the control and RSV model groups were treated intragastrically with 0. 9% normal saline and those in the Ribavirin and QFOL groups with Ribavirin at 0. 0025 g/m L and QFOL at 1. 33 g/m L and 4 g/m L,respectively,all for 72 hours. Then all the mice were killed and the lung tissue harvested from 5 animals in each group for pathological analysis,while the levels of IL-10 and IL-17 in the bronchoalveolar lavage fluid of the other 5 detected by ELISA. The expressions of the cytokines Treg and Th17＇ in the spleen from 4 mice in each group were determined by flow cytometry. Results Compared with the RSV models,pathologic changes were significantly reduced in the mice of the QFOL,Ribavirin and control groups（ P〈0. 01）,the expression of IL-10 remarkably up-regulated in the lowdose QFOL,high-dose QFOL,Ribavirin,and control groups（ [39. 21 ± 1. 57]vs [43. 54 ± 1. 03],[46. 64 ± 0. 48],[47. 83 ± 0. 87],and [50. 44 ± 1. 04] ng/L,all P〈0. 01）,while the level of IL-17 markedly down-regulated（ [70. 96 ± 0. 53] vs [55. 92 ± 0. 83],[33. 66 ±0. 70],[21. 92 ±1. 38],and [9. 42 ± 0. 59] pg/m L,all P〈0. 01）. The expressions of Treg/Th17＇ were significantly increased in the low-dose QFOL,high-dose QFOL,Ribavirin,and control groups（ 2. 89 ± 0. 52,6. 38 ± 0. 36,3. 95 ± 0. 26,and 3. 54 ±0. 85） as compared with that in the RSV models（ 0. 96 ± 0. 16）（ all P〈0. 01）. Both low- and high-dose QFOL groups showed statistically significant differences from the Ribavirin group in the levels of Treg,Th17,and Treg/Th17（ P〈0. 05）. Conclusion QFOL can regulate the balance of Treg/Th17,increase the expression of IL-10 and decrease that of IL-17 in the lung tissue of RSV-infected mice,which further proves the efficacy of QFOL in the treatment of RSV-induced pneumonia.