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目的探讨气道应用T-bet重组腺病毒(AdT-bet)对哮喘小鼠气道炎症反应的影响及机制。方法C57BL/6小鼠随机分为4组,实验组(A组)、对照病毒组(B组)、PBS对照组(C组)和正常对照组(D组),A、B、C组采用卵蛋白(OVA)、明矾建立哮喘模型,分别于第19天激发前气道内单次应用50μLAdT-bet(108pfu)、AdLacZ、PBS。26d取肺泡灌洗液(BALF)测定细胞成分、IL-4、IL-5、IFNγ浓度,取血测定血浆IgE水平,观察肺组织病理学变化及GATA-3表达。结果1)A组BALF中的嗜酸粒细胞(EOS)为(0.5±0.2)%明显低于B组(21.2±6.9)%和C组(20.9±6.8)%(P<0.01);2)A组BALF中的IL-4(6.7±3.8)pg/mL和IL-5(12.4±4.9)pg/mL的水平明显低于B组[IL-4(91.4±22.5)pg/mL和IL-5(55.6±10.6)pg/mL]和C组[IL-4(89.8±23.6)pg/mL和IL-5(56.7±11.5)pg/mL](P<0.01),而IFNγ的水平(710±45)pg/mL则明显高于B组(13.1±3.5)pg/mL和C组(11.9±4.8)pg/mL;3)A组血浆中总IgE(39.1±9.6)pg/mL和OVA特异性IgE(18.8±4.9)U/mL水平显著低于B组[总IgE(67.1±9.9mg/mL),OVA特异性IgE(34.5±7.7)U/mL]和C组[总IgE(68.4±9.8)mg/mL,OVA特异性IgE(35.7±8.3)U/mL](P<0.01);4)HE染色示B组和C组支气管平滑肌肥厚,黏膜充血、水肿,黏膜层增厚,并有EOS为主的炎性细胞浸润,管腔内可见黏液栓,支气管壁周围有EOS为主的炎性细胞浸润,而A组小鼠的上述炎症性改变则明显减轻;5)免疫组化显示B组和C组肺组织中GATA-3的表达明显增加,而A组GATA-3的表达明显减少。结论本研究表明气道应用AdT-bet可抑制哮喘鼠IL-4、IL-5合成,增强IFNγ表达,抑制EOS在气道内的炎性浸润及血浆IgE水平,机制可能与AdT-bet增强Th1反应,抑制GATA-3表达,从而抑制Th2型免疫反应有关。
Objective To investigate the effect of airway T-bet recombinant adenovirus (AdT-bet) on airway inflammation in asthmatic mice and its mechanism. Methods C57BL / 6 mice were randomly divided into 4 groups: experimental group (group A), control virus group (group B), PBS control group (group C) and normal control group (group D) (OVA) and alum to establish a model of asthma. Fifty microliters of AdT-bet (108 pfu), AdLacZ and PBS were given to the airway before the stimulation on the 19th day. The levels of IL-4, IL-5 and IFNγ were measured by BALF after 26 days. The levels of plasma IgE were determined by blood sampling. The pathological changes and the expression of GATA-3 were observed. Results (1) The eosinophilic granulocytes (EOS) in group A were significantly lower than that in group B (21.2 ± 6.9)% and group C (20.9 ± 6.8)% (P <0.01) The levels of IL-4 (6.7 ± 3.8) pg / mL and IL-5 (12.4 ± 4.9) pg / mL in BALF in group A were significantly lower than those in group B [91.4 ± 22.5 pg / mL and IL- 5 (55.6 ± 10.6) pg / mL] and group C (IL-4 89.8 ± 23.6 pg / mL and IL-5 56.7 ± 11.5 pg / mL] ± 45) pg / mL was significantly higher than those in group B (13.1 ± 3.5) pg / mL and group C (11.9 ± 4.8) pg / mL; The level of specific IgE (18.8 ± 4.9) U / mL was significantly lower than that of group B [total IgE (67.1 ± 9.9 mg / mL) and OVA specific IgE (34.5 ± 7.7) U / mL] (P <0.01); 4) HE staining showed bronchial smooth muscle hypertrophy, mucosal congestion, edema, thickening of mucosa in group B and group C, Inflammatory cells infiltrated mainly in EOS. Mucus suppositories were found in the lumen. Inflammatory cells infiltrated mainly around the bronchial wall were observed in EOS group. The inflammatory changes in group A were significantly alleviated. 5) Immunohistochemistry The results showed that the expression of GATA-3 in group B and group C was significantly increased, while the expression of GATA-3 in group A was significantly decreased. Conclusions This study shows that airway AdT-bet can inhibit the synthesis of IL-4 and IL-5 in asthmatic mice, enhance the expression of IFNγ, inhibit inflammatory infiltration of EOS in airway and plasma IgE level, which may be related to the increase of Th1 response to AdT-bet , Inhibit GATA-3 expression, thereby inhibiting the Th2-type immune response.