下呼吸道流感嗜血杆菌定植对哮喘小鼠气道炎症的影响及信号通路的研究

doi:10.16150/j.1671-2870.2020.01.010

摘要/Abstract

摘要：

目的：观察下呼吸道流感嗜血杆菌定植对哮喘小鼠气道炎症的影响,并研究其信号通路。方法：采用C57/B6和TLR4^-/-小鼠,分别用卵清蛋白（ovalbumin,OVA）致敏和激发制备慢性哮喘小鼠模型,用流感嗜血杆菌琼脂菌珠制作气道定植模型（注菌组为2组,注菌后第7天和第21天组）,同时设置对应时间（第7天和第21天）的0.9%NaCl溶液注射组及单纯流感定植组、单纯哮喘组,每种小鼠各8组,共16组。在哮喘小鼠气道注菌之后的第7、21天处死小鼠,检测血清中肿瘤坏死因子α（tumor necrosis factor α,TNF-α）含量,采用免疫组织化学法检测小鼠肺组织髓样分化因子88（myeloid differentiation factor 88, MyD88）和核因子κB（nuclear factor κB, NF-κB）的表达,并分别与相同时间的单纯流感定植及0.9%NaCl溶液注射、单纯哮喘对照小鼠相比较;再将C57/B6与TLR4^-/-小鼠的实验结果进行比较,同时比较各组小鼠支气管肺泡灌洗液（bronchoalveolar lavage fluid,BALF）中的细胞总数。结果：与相同时间的单纯流感定植及0.9%NaCl溶液注射的对照小鼠相比,有流感嗜血杆菌呼吸道定植的C57/B6和TLR4^-/-哮喘小鼠血清中TNF-α含量、肺组织中MyD88和NF-κB的表达以及BALF中的细胞总数均升高（P均<0.05）;与同时间对应组C57/B6小鼠相比,合并流感嗜血杆菌呼吸道定植的TLR4^-/-哮喘小鼠血清中TNF-α含量、肺组织中MyD88和NF-κB的表达以及BALF中的细胞总数均降低P均<0.05）。结论：流感嗜血杆菌在下呼吸道定植可以通过激活TLR4-MyD88通路,促进转录因子NF-κB的表达,加重哮喘的气道炎症,这可能是哮喘发病的重要机制之一。

关键词: 流感嗜血杆菌, 哮喘, 气道炎症, 信号通路

Abstract:

Objective: To observe the effect of Haemophilus influenzae colonizing lower respiratory tract on airway inflammation in asthmatic mice and study the related signaling pathway. Methods: Thirty-two C57/B6 mice were sensitized and challenged with ovalbumin (OVA) to establish a mice model of chronic asthma (AC group), and then half of them received intratracheal injection of Haemophilus influenza coated in agar beads to built a mice model of chronic asthma with airway Haemophilus influenza colonization (AS group). Besides, 16 C57/B6 mice were treated with intratracheal injection of Haemophilus influenza (NS group) and 16 C57/B6 mice received intratracheal injection of 0.9% NaCl (NC group). Sixty-four TLR4^-/- mice were treated and grouped as C57/B6 mice. Mice including C57/B6 and TLR4^-/- mice were sacrificed at 7 and 14 days after intratracheal injection, and serum tumor necrosis factor α(TNF-α) content, total number of cells in bronchoalveolar lavage fluid( BALF), and the expression of MyD88 and NF-κB in lung tissue were measured. The indice in AS group were compared with those in the control groups (AC, NS and NC) at corresponding time. Results: The serum level of TNF-α, total number of cells in BALF, and expression of MyD88 and NF-κB in lung tissue were higher in AS group than those in NC and NS group (both in C57/B6 mice and TLR4^-/- mice). Compared with C57/B6 wild-type mice, TLR4^-/- mice in AS group had lower levels of serum TNF-α, decreased numbers of cells in BALF, and down-regulated expression of MyD88 and NF-κB in lung tissue. Conclusions: Lower airway colonization with Haemophilus influenzae may aggravate airway inflammation of asthma by activating the TLR4-MyD88 pathway and promoting expression of NF-κB, which may be one of the important mechanisms of asthma.

Key words: Haemophilus influenza, Asthma, Airway inflammation, Signaling pathway

中图分类号:

R563.25

康建强, 董杨阳, 杨玲, 宋珍, 范嘉盈. 下呼吸道流感嗜血杆菌定植对哮喘小鼠气道炎症的影响及信号通路的研究[J]. 诊断学理论与实践, 2020, 19(1): 44-49.

KANG Jianqiang, DONG Yangyang, YANG Ling, SONG Zhen, FAN Jiaying. Effect of Haemophilus influenzae colonizing lower respiratory tract on airway inflammation and its signaling pathway in asthmatic mice[J]. Journal of Diagnostics Concepts & Practice, 2020, 19(1): 44-49.

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分组	第7天		第21天
分组	C57/B6组	TLR4^-/-组	C57/B6	TLR4^-/-组
NC	29.00±2.64	31.00±3.64	28.69±1.71	29.33±126
NS	200.67±9.50^a)	109.35±10.36^a),c)	100.67±9.5^a)	53.10±2.04^a),c)
AC	120.67±20^a)	130.76±8.2^a)	119.67±9.9^a)	129.02±10.1^a)
AS	313.67±6.4^a),b)	200.61±9.52^a),b),c)	151.67±14.4^a)	137.3±7.53^a)

分组	第7天			第21天
分组	C57/B6组	TLR4^-/-组	C57/B6组	TLR4^-/-组
NC	10.83±1.96	11.60±8.70	12.49±6.72	11.60±0.17
NS	14.61±3.59^a)	15.89±1.41	19.57±5.25	18.58±1.79
AC	14.46±3.65	18.02±4.23^a)	14.47±3.84	17.64±5.78
AS	150.59±66.94^a),b)	28.26±5.22^a),b),c)	76.09±22.64^a),b)	38.98±3.13^a),b),c)