摘要
目的建立重组屋尘螨Derp2变应原(rDerp2)诱导的小鼠变态反应气道炎症动物模型。方法在大肠杆菌中诱导表达Derp2重组蛋白,用镍亲和柱层析法提纯重组蛋白;32只BALB/c小鼠随机分为屋尘螨粗浸液组(A组)、屋尘螨粗浸液+rDerp2组(B组)、rDerp2组(C组)、对照组(D组)。分别观察肺组织病理变化与支气管肺泡灌洗液(BLAF)中细胞学变化,采用酶联免疫吸附试验(ELISA)测定BLAF中和脾细胞培养上清IL-4与INF-γ变化,ELISA测定血清中IgE、IgG1与IgG2a抗体变化。结果成功表达、纯化Derp2重组蛋白;A、B、C组肺部病理改变呈现明显的变态反应性炎症;A、B、C组小鼠BALF中的细胞总数、淋巴细胞、中性粒细胞数和EOS计数显著高于D组(P<0.01);A、B、C组BALF中IL-4含量分别为(109.35±16.46)pg/ml、(88.87±5.66)pg/ml、(85.59±6.05)pg/ml,与D组(23.17±2.67)pg/ml相比差异有统计学意义(P均<0.01)。A、B、C组抗原特异性IgE抗体分别为0.49±0.04、0.41±0.02、0.37±0.04,与D组(0.03±0.01)相比差异有统计学意义(P均<0.01);A、B、C组小鼠脾细胞分泌IL-4分别为(266.20±19.18)pg/ml、(252.72±15.81)pg/ml、(243.62±19.07)pg/ml,与D组(15.99±1.56)pg/ml相比差异有统计学意义(P均<0.01)。结论用rDerp2能成功建立小鼠变态反应气道炎症动物模型。
Objective To develop a mouse model of allergic airway inflammation with recombinant house dust mite allergen （Derp2） as an allergen. Methods The recombinant Derp2 was expressed in Escherichia coli and purified through Ni^＋ NTA His agarose. Thirty two BALB/c mice were randomly divided into crude mite extract group（group A, n = 8）, rDerp2 ＋ crude mite extract group（group B, n = 8）, rDerp2 groups（group C, n = 8） and control groups（group D, n = 8）. The mice were intranasally challenged after sensitization. Lungs were fixed in situ, processed and stained with haematoxylin and eosin. Differential cell counting of bronchoalveolar lavage fluid（BALF） was used to as indicator airway of inflammation. IL-4/INF-γ in BALF and antibody responses of IgE, IgG1 and IgG2a in serum were assessed with EIJSA. Cells from spleen were cultured for 24 h with rDerp2, and IL- 4/INF-γ in supernatants was measured by ELISA. Results Mice in A, B and C group developed eosinophil infiltration of the airways. Total cells and eosinophils in BALF were induced in A, B and C group. IL-4 in BALF in A （ 109.35 ± 16.46） pg/ml, B （88.87 ± 5.66） pg/ml and C （ 85.59 ± 6.05） pg/ml group were significantly different from those in D group [（23.17±2.67） pg/ml, P〈0.01]. IgE antibody in A（0.49± 0.04）, B（0.41 ±0.02） and C （0.37 ± 0.04） group were significantly different from those in group D [ （0.03 ± 0.01 ）, P 〈 0.01 ]; IL-4 released from spleen cultured into supernatants in A（266.20± 19.18） pg/ml, B （252.72± 15.81） pg/ml and C （243.62 ± 19.07） pg/ml group were significantly different from those in group D [ （15.99 ± 1.56） pg/ml, P 〈 0.01 ]. Conclusion The mouse model can be applied in the investigation of the immunopathogenesis of human asthma.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2005年第7期564-569,共6页
Chinese Journal of Microbiology and Immunology
基金
国家863计划(No.2002AA214011)
国家自然科学基金(No.30271226
30260101)
