血管内皮生长因子受体抑制剂对小鼠支气管哮喘模型气道炎症和气道重塑的影响

The effect of a vascular endothelial growth factor receptor inhibitor on airway inflammation and airway remodeling in a murine model of asthma

目的研究血管内皮生长因子(VEGF)受体抑制剂对变应性气道炎症和气道重塑的影响,阐明VEGF与支气管哮喘(简称哮喘)以及气道重塑的关系。方法BALB/c小鼠按随机数字表法分为正常对照组(A组)、哮喘模型组(B组)、VEGF受体抑制剂治疗组(C组),每组各10只。用酶联免疫吸附测定(ELISA)法对各组小鼠支气管肺泡灌洗液(BALF)和血清中VEGF进行定量分析;用免疫组化法检测VEGF在小鼠肺组织内的表达水平。采用医学图像分析软件测定支气管管壁厚度(WAt/P i)、支气管平滑肌厚度(WAm/P i)、支气管平滑肌细胞计数(N/P i)及肺组织切片中的血管计数、血管壁平滑肌厚度、血管壁平滑肌细胞计数。结果BALF中细胞总数和嗜酸粒细胞比值B组分别为(142±63)×107/L、98.0±46.9,A组分别为(30±14)×107/L、0.7±1.1,C组分别为(41±17)×107/L、4.9±3.5,A组和C组BALF中细胞总数和嗜酸粒细胞比值分别与B组比较差异有统计学意义(P均<0.01)。B组BALF中上清液和血清中VEGF水平分别为(55±26)pg/m l、(72±26)pg/m l,A组分别为(37±9)pg/m l、(49±18)pg/m l,C组分别为(34±3)pg/m l、(43±19)pg/m l,A组与B组、C组与B组间比较差异均有统计学意义(P均<0.05)。免疫组化结果显示,B组大部分支气管平滑肌、黏膜上皮细胞、肺泡上皮细胞和血管周围VEGF均呈阳性表达,而A组和C组几乎没有VEGF的表达。图像分析显示,B组WAt/P i、WAm/P i、N/P i分别为(17±5)μm2/μm、(6.3±2.2)μm2/μm、(0.050±0.020)个/μm,A组分别为(8±3)μm2/μm、(3.2±0.8)μm2/μm、(0.027±0.017)个/μm,A组与B组比较差异有统计学意义(P分别<0.01、0.05)。B组和A组血管计数分别为(19±3)个、(10±5)个,A组与B组比较差异均有统计学意义(P<0.01)。经抑制剂治疗后C组WAm/P i、血管计数分别为(4.5±1.3)μm2/μm、(11±3)个,C组与B组比较差异均有统计学意义(P均<0.05)。结论VEGF在小鼠哮喘模型气道及肺内过度表达,并参与了哮喘的发病和气道重塑过程。VEGF受体抑制剂可明显改善哮喘小鼠的变应性气道炎症和气道重塑的病理生理过程。

Objective To investigate the expression of vascular endothelial growth factor（VEGF） and the effect of a VEGF receptor inhibitor on airway inflammation and remodeling in a murine model of asthma. Methods BALB/c mice were randomly divided into three groups： group A （control group, n = 10）, group B（asthmatic model group, n = 10） and group C（VEGF receptor inhibitor group, n = 10）. The asthmatic model was established by OVA sensitization and challenge. Bronchoalveolar lavage fluid（BALF） cell count and differential were carried out, VEGF levels in BALF and serum were measured by enzyme- linked-immunosorbent assay（ ELISA）, the pathologic changes of bronchi and the lung tissue were evaluated, and the expression of VEGF was analyzed with immunohistochemistry （ISH）. The airway wall thickness （WAt/Pi）, the bronchial smooth muscle thickness （WAm/Pi）, the bronchial smooth nucleus counts （N/Pi） and vascular counts were measured by using image analysis system. Results There was a significant increase in the numbers of total cells and eosinophils（EOS） in the BALF of group B[ （ 142 ±63） × 10^7/L,98.0 ± 46. 9 compared to those of group A （ 30 ± 14 ） × 10^7/L, 0. 7 ± 1.1, all P 〈 0. 01 ]. After administration of the VEGF receptor inhibitor, the numbers of total cells and EOS were significantly decreased [（41 ±17） × 10^7/L, 4.9 ±3.5, all P〈0.01]. BALF and serum levels of VEGF in group B were [ （55 ±26） pg/ml, and （72 ±26） pg/ml] respectively. They were significantly higher than those in group A [ （37±9）pg/ml, （49 ± 18）pg/ml, respectively] and in group C[（34 ±3） pg/ml, （43 ± 19） pg/ml, respectively]. ISH showed that expression of VEGF in group B was increased compared with group A and C. WAt/Pi, WAm/Pi, and N/Pi in group B [ （ 17 ± 5 ） μ m^2/μm, （6.3 ± 2. 2 ）μm^2/μm , （0. 050 ± 0. 020）/μm, respectively ] were all significantly higher than those in group A [ （8 ± 3 ） μm^2/μm, （3.2 ± 0. 8） μm^2/μm, （ 0. 027 ± 0. 017 ）/μm respectively, P 〈 0. 01 and P 〈 0. 05 ]. The vascular count of the airways in group B （ 19 ± 3 ） was significantly increased compared to that of group A （ 10 ± 5, P 〈 0. 01 ）. Administration of the VEGF receptor inhibitor reduced WAm/Pi （4. 5 ± 1.3 ）μm^2/μm, P 〈0. 05 ） and the vascular count （11 ± 3, P 〈 0. 01 ）. Conclusions VEGF was over-expressed in the murine model of asthma, and involved in angiogenesis and airway remodeling. VEGF receptor inhibitors may be effective in reducing allergic airway inflammation and airway remodeling.