施焕中

论文题目：白细胞介素—５在支气管哮喘发病机理中的作用

作者简介：施焕中，男，1964年10月出生，1996年09月师从于广西医科大学刘义教授，于2000年6月获博士学位。

摘  要

 

目的  (1)了解过敏性支气管哮喘患儿血清白细胞介素-5(IL-5)的水平、来源及其意义；(2)研究IL-5对哮喘患者气道嗜酸性粒细胞(EOS)性炎症的影响; (3)探讨IL-5在气道高反应性发生机制中的作用; (4)阐明IL-5外周血嗜酸性粒细胞数及其活化状态的影响；⑸探讨IL-5增加气道可溶性细胞间粘附分子-1(sICAM-1)水平的机制。最终系统阐明IL-5在支气管哮喘发病机理中的作用。

方法  (1) 测定过敏性哮喘患儿血清IL-5水平,并分析其与嗜酸阳离子蛋白(ECP)及可溶性IL-2受体(sIL-2R)之间的关系。(2) 将人重组IL-5经纤维支气管镜直接注入哮喘患者（8例）右中叶或左舌叶的肺段支气管，而将载体溶液注入对侧肺段支气管作为对照。24小时后复经纤支镜于相应的肺段支气管收集支气管肺泡灌洗液（BALF）及支气管粘膜组织，对BALF细胞进行计数和分类，测定其中ECP水平，并以免疫组织化学技术显示粘膜组织中总EOS（BMK-13⁺细胞）及活化EOS（EG2⁺细胞）的浸润情况。(3) 另随机选择8例哮喘患者雾化吸入人重组IL-5，并分别于吸入前、吸入后2小时、24小时和48小时测定其气道反应性（以PC₂₀-Mch表示）及计数痰液中EOS数。(4) 于哮喘患者吸入IL-5前后进行外周血有核细胞计数和分类，并以放射免疫法测定血清中ECP及总IgE水平。(5) 观察哮喘患者吸入IL-5后痰和血清sICAM-1水平的变化，并与正常人作对照研究，通过与白蛋白含量对比，分析气道所增加的sICAM-1的来源。

结果  (1)急性发作期哮喘患儿血清IL-5、ECP和sIL-2R水平均明显高于缓解期患者和正常对照组；而且缓解期患者和正常对照组除sIL-2外，其差异也十分明显；ECP水平和IL-5水平之间以及IL-5水平和sIL-2R水平呈明显的正相关关系。(2)直接以IL-5刺激肺段支气管可以引起BALF中EOS数和ECP水平显著增高。而且，IL-5刺激组支气管粘膜BMK13⁺和EG2⁺细胞数也明显高于对照组。(3)哮喘患者吸入IL-5后PC₂₀-Mch随时间而明显降低，至24小时达最低值，48小时后仍维持在较低水平；这种PC20-Mch的降低伴随着痰EOS数的相应增高。(4)哮喘患者吸入IL-5后外周血中EOS占细胞总数的百分比、EOS绝对细胞数以及ECP水平均随时间而明显升高，至24小时达最高值，48小时后仍维持在较高水平；而IL-5在整个实验过程中对血清总IgE水平并无明显影响。⑸哮喘患者吸入IL-5后24小时sICAM-1即明显高于吸入前基础值；48小时后sICAM-1水平达最大值，而这种高水平的sICAM-1至少可以维持72小时。我们还注意到，对于正常人而言，即使给予IL-5刺激亦不能引起其痰sICAM-1水平出现任何变化。和痰sICAM-1变化情况相类似，吸入IL-5可以明显地招致哮喘患者血清sICAM-1水平随着时间而显著增高。此外，哮喘患者吸入IL-5后实际增加的痰液sICAM-1远远高于其预计增加值。

结论  (1) IL-5参与了哮喘急性发作过程, 且其含量升高与病情严重程度有一定的关系，还可能与哮喘气道慢性炎症持续存在的病理生理过程有关。血清中IL-5可能来自激活的T淋巴细胞。(2) IL-5不但可以趋化EOS润到哮喘患者的气道，还可促使其活化从而释放炎症介质。(3) IL-5可以促使哮喘患者气道反应性明显增高，而IL-5导致气道高反应性很可能是通过EOS的作用而实现。(4) 吸入IL-5不仅可以促使循环中EOS数明显增多，而且还能导致其活化从而参与哮喘的发病。(5) 哮喘患者吸入IL-5后痰液和血清sICAM-1水平显著增高，其升高机制很可能是IL-5刺激气道局部产生sICAM-1所致，而不可能仅仅是由于血清sICAM-1渗漏到气道的缘故。既然sICAM-1参与哮喘的气道EOS浸润过程和气道高反应性的发生和发展，而IL-5能提高sICAM-1的水平，推测IL-5加重哮喘患者气道嗜酸细胞性炎症和促使气道反应性增高的一个重要机制很可能就是依赖sICAM-1的作用而实现的。

 

Abstract

 

Objective  (1) To investigate the concentration, source and significance of serum interleukin-5 (IL-5) in children with allergic asthma. (2) To investigate the effect of IL-5 on eosinophilic inflammation in asthmatic airways in vivo. (3) To investigate the role of IL-5 in the mechanism of airway hyperresponsiveness. (4) To investigate the effects of IL-5 on changes of activity and number of circulating eosinophils, as well as concentrations of serum IgE. (5) To investigate the effects of IL-5 on changes of soluble intracellular adhesion molecule-1 (sICAM-1) concentrations in induced sputum from allergic asthmatics.

Methods  (1) Sera were collected from children with allergic asthma, including both acute cases and remission cases, and health subjects. ELISA assay was adopted to detect the concentrations of IL 5 and soluble IL-2 receptor (sIL-2R), and immunoradioassay was used to detect the levels of eosinophil cationic protein (ECP). (2) Eight subjects with mild atopic asthma underwent initial bronchoscopy during which control bronchoalveolar lavage fluid (BALF) as well as bronchial mucosa were obtained, and at the same time, normal saline and IL-5 were administered to two sublobar segments separately. The second bronchoscopy was carried out and the samples from challenged sites were taken 24 h later. (3) The effects of inhaled IL-5 on airway responsiveness to methacholine and cell populations in induced sputum were studied in eight patients with allergic bronchial asthma using a placebo-controlled study design. (4) Eight nonsmoking patients with allergic asthma were administered IL-5 by nebulization, total blood nuclear cell counts and differentials, as well as concentrations of ECP and IgE in serum were determined before, and at 2, 24, 48 h after inhalation, respectively. (5) Eight patients with allergic asthma and six non-allergic normal subjects were administered IL-5 by nebulization, and the concentrations of sICAM-1 in induced sputum from each subject were determined before, and at 2, 24, 48, 72 h after inhalation, respectively.

Results  (1) The levels of IL-5, ECP and sIL-2R were significantly raised during asthma attack when compared with those observed in normal controls; IL-5 and ECP levels were also significantly higher during asthma attack than those observed in stable asthma, with the difference of sIL-2R did not reach significant. In addition, the elevation of IL-5 was related to concentrations of ECP and sIL 2R. (2) The "total" eosinophils (BMK-13⁺ cells) and the "activated" eosinophils (EG2⁺ cells) in bronchial mucosa, the eosinophil numbers in BAL fluid, as well as ECP in BAL fluid from saline challenged segments were not different from those in unchallenged segments. However, a significant eosinophilia was observed in bronchial mucosa and BALF from IL-5 challenged-sites. Eosinophil activation, as assessed by secretion of ECP, was also increased significantly in bronchial mucosa and BAL fluid. (3) The inhalation of IL-5 did not alter lung function in allergic asthmatics. In the control experiments receiving vehicle, PC₂₀-Mch values did not change nor did the numbers of eosinophils or ECP sputum values change from baseline. In contrast, after IL-5 inhalation, PC₂₀-Mch fell down significantly. Accompanying this increased airway sensitivity was a significant eosinophilia and elevated concentrations of eosinophil cationic protein in induced sputum. (4) Eosinophil numbers in circulation increased after IL-5 inhalation, accompanying this significant blood eosinophilia was a significant elevation of serum ECP levels. However, IL-5 inhalation had no significant effect of levels of serum total IgE. (5) In allergic asthmatics, vehicle challenge was not able to cause any changes in sputum sICAM-1 concentrations. However, there were increases in sputum sICAM-1 concentrations after IL-5 inhalation, which increased with time significantly greater than baseline values, reaching a maximum at 48 h, and lasting no less than 72 h. The concentrations of sICAM-1 in sputum after IL-5 challenge exceeded levels that could be accounted for by passive transudation from the circulation, based upon the magnitude of increases in sputum albumin concentration.

Conclusions  (1) IL-5 is involved in the acute exacerbation of asthma, and may be related to chronic inflammation in the asthmatic airways. Our results suggested that IL-5 derived from activated lymphocytes. (2) IL-5 is capable of inducing eosinophil infiltration into the asthmatic airways, as well as the activation of infiltrating eosinophils. (3) IL-5 increases airway responsiveness and infiltration of activated eosinophils into the airway in patients with allergic bronchial asthma. (4) IL-5 not only induced a significant blood eosinophilia, but also resulted in the activation of circulating eosinophils. (5) IL-5 inhalation was capable of inducing an elevation of sputum sICAM-1 concentration by stimulating its local release in allergic asthmatics, but not in non-allergic normal subjects.