支气管哮喘大鼠气道重塑中肺泡巨噬细胞的作用

The role of alveolar macrophages in airway remodeling in asthmatic rats

目的研究支气管哮喘(简称哮喘)大鼠气道重塑过程中肺泡巨噬细胞(AM)的变化及其作用。方法48只清洁级雄性幼年SD大鼠按随机数字表法分为正常对照组(A组)、哮喘3d组(B组)、哮喘14d组(C组)和哮喘30d组(D组),每组12只。应用鸡卵白蛋白(OVA)建立哮喘大鼠模型,纯化支气管肺泡灌洗液(BALF)中的AM,测定AM中肿瘤坏死因子α(TNF-α)、前列腺素E2(PGE2)的含量和基质金属蛋白酶9基因(MMP-9mRNA)及其组织抑制物1基因(TIMP-1mRNA)的表达,并测量大鼠支气管壁总面积和平滑肌面积,计算单位基底膜周径(Pbm)的支气管壁厚度(WAt)和平滑肌厚度(WAm)。结果D组WAt和WAm[(85±9)μm2/μm、(28.6±4.9)μm2/μm]与A组[(67±10)μm2/μm、(16.8±2.4)μm2/μm]比较差异有统计学意义(t值分别为2.938、3.227,P均<0.01);D组AM中TNF-α和PGE2含量[(0.68±0.25)μg/L、(0.122±0.030)μg/L]与A组[(0.37±0.09)μg/L、(0.079±0.018)μg/L]比较差异有统计学意义(t值分别为2.683、3.016,P均<0.01),与B组[(0.74±0.29)μg/L、(0.120±0.028)μg/L]、C组[(0.71±0.23)μg/L、(0.117±0.028)μg/L]比较差异无统计学意义(t值分别为1.624、0.472、0.935、0.533,P均>0.05);D组AM中MMP-9mRNA及TIMP-1mRNA含量吸光度(A)值分别为0.346±0.033、0.361±0.040,与C组(0.279±0.015、0.259±0.015)比较差异有统计学意义(t值分别为2.574、2.716,P均<0.01),D组(0.183±0.025)与B组(0.136±0.014)比较差异有统计学意义(t值分别为2.913、3.017,P均<0.01),D组(0.104±0.007)与A组(0.109±0.008)比较差异有统计学意义(t值分别为3.632、3.487,P均<0.01);各组AM中MMP-9mRNA含量与WAt、WAm呈正相关(r值分别为0.693、0.738,P均<0.01),AM中TIMP-1mRNA含量与WAt、WAm呈正相关(r值分别为0.823、0.876,P均<0.01)。结论哮喘大鼠AM及其分泌的一些细胞因子与气道重塑关系密切。

Objective To study the role of alveolar macrophages （AM） in the processes of airway remodeling in asthmatic rats. Methods Forty-eight young male Sprague-Dawley rats （ Grade Ⅱ ） were divided randomly into a control group （A group）, a 3 day asthma group （B group）, a 14 day asthma group （ C group） and a 30 day asthma group （ D group）. The rats were sensitized and challenged by ovalbumin to establish the asthmatic model. AM were purified from bronchoalveolar lavage. The content of tumor necrosis factor-α （TNF-α） in AM was measured by enzyme-linked immunosorbent assay, prostaglandin E2 （PGE2 ） by radioimmunoassay, matrix metalloproteinase-9 mRNA （MMP-9 mRNA ） and tissue inhibitor of metalloproteinase-1 mRNA （TIMP-1 mRNA） by hybridization in situ. The total bronchial wall area （WAt） and the smooth muscle area （WAm） were measured by image analysis system. The WAr and the WAm were quantified per unit length of basement membrane （Pbm）. Results The bronchial wall thickness and the smooth muscle thickness of D group [ （85 ±9） μ m^2/μm, （28.6 ±4. 9） μ m^2/μm] were significantly higher than those of A group [ （67 ± 10） μm^2/μm （ 16. 8 ±2. 4） μm^2/μm,t =2. 938,3. 227, all P 〈0. 01 ]. The contents of TNF-α and PGE2 in D group [ （0. 68 ±0.25） g/L, （0. 122 ±0. 030） μg/L] were significantly higher than those in A group [（0.37 ±0.09） μg/L, （0.079 ±0.018） μg/L,t =2.683, 3.016, all P 〈0. 01 ]. When A group was compared with B group [ （0. 74 ± 0. 29 ）μL, （0. 120 ± 0. 028 ） μg/L ] and C group [ （0. 71 ± 0. 23 ） μg/L, （ 0. 117 ± 0. 028 ） μg/L ] , there was no significant difference （ t = 1. 624, 0. 472, all P 〉 0.05 ） and （ t = 0. 935,0. 533, all P 〉 0. 05 ）. The contents （ light density） of MMP-9 mRNA and TIMP-1 mRNA in D group （0. 346 ± 0. 033, 0. 361 ± 0. 040） were significantly higher than C group （0. 279 ± 0. 015, 0. 259 ± 0. 015, t = 2. 574,2. 716, all P 〈 0. 01 ）, and so did D group and B group （0. 183 ±0. 025, 0. 136 ±0. 014,t = 2. 913,3.017, all P 〈0. 01 ）, D group and A group（0. 104 ±0. 007, 0. 109 ±0. 008 ,t =3. 632,3.487, all P 〈0. 01 ）. There were significant correlations between the contents of MMP-9 mRNA and WAt （r = 0. 693, P 〈 0.01）, between MMP-9 mRNA and WArn （r = 0.738, P 〈 0. 01 ）, between TIMP-1 mRNA and WAt （ r = 0. 823, P 〈 0. 01 ）, and between TIMP-1 mRNA and WAm （r = 0. 876, P 〈 0.01 ）. Conclusion AM and AM-derived cytokines are associated with airway remodelingin asthmatic rats.