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目的探讨白细胞介素-12(IL-12)在哮喘小鼠发病机制中的作用及对气道炎症的影响。方法 65只小鼠随机分为对照组(A组)、哮喘组(B组)、重组白细胞介素-12(rm IL-12)组(C组)和IL-12基因敲除组(D组),以卵白蛋白(OVA)致敏和激发制备哮喘模型。C组在每次激发前30 min给予rm IL-12腹腔注射,D组在激发前1 d给小鼠植入含有抗IL-12多克隆抗体的缓释泵,A组小鼠在末次激发后24 h处死,其余三组小鼠在末次激发后的1 d、2 d、4 d、7 d分别处死。摘眼球取血进行血浆一氧化氮(NO)和免疫球蛋白E(Ig E)的检测。收集各组小鼠的支气管肺泡灌洗液(BALF),ELISA法检测其上清中白细胞介素-10(IL-10)、IL-12、肿瘤坏死因子-α(TNF-α)含量。离心后的细胞沉淀行白细胞总数和嗜酸性粒细胞(EOS)分类计数;剪下结扎右肺下叶用于HE染色。结果(1)细胞计数:小鼠BALF中的细胞总数和EOS百分比,B组明显高于A组(P<0.05),C组明显低于B组(P<0.05),D组较B组明显升高(P<0.05)。(2)ELISA检测:与A组相比,B组小鼠BALF中IL-10、IL-12水平明显降低(P<0.05),TNF-α、NO、Ig E水平明显升高(P<0.05)。与B组相比,C组小鼠BALF中IL-10、IL-12水平明显升高,TNF-α、NO、Ig E水平明显降低;D组小鼠TNF-α、NO、Ig E水平明显升高,IL-10、IL-12水平明显降低;于1 d、2 d、4 d、7 d差异均有统计学意义(P<0.05)。(3)肺组织病理改变:A组小鼠HE染色未见异常;B组小鼠肺泡间隔及小血管周围可见大量炎性细胞浸润;C组小鼠的炎性细胞浸润较B组缓解;D组小鼠部分可见小气道管腔完全闭塞,炎性细胞浸润较B组明显加重。结论 rm IL-12能够明显抑制哮喘小鼠炎症细胞的浸润,同时降低炎症因子TNF-α、NO和Ig E的表达,促进抑炎因子IL-10的表达,从而在哮喘发病过程中起到保护作用。
Objective To investigate the role of interleukin-12 (IL-12) in the pathogenesis of asthma in mice and its effect on airway inflammation. Methods Sixty-five mice were randomly divided into control group (group A), asthma group (group B), recombinant interleukin-12 (group C) and IL-12 knockout group ), Sensitized and challenged with ovalbumin (OVA) to prepare an asthma model. The mice in group C were injected intraperitoneally with rm IL-12 at 30 min before each challenge, and the mice in group D were injected with anti-IL-12 polyclonal antibody one day before challenge. 24 h after sacrifice, and the other three groups were sacrificed on the 1st, 2nd, 4th and 7th days after the last challenge. Eyedrops were taken for detection of plasma nitric oxide (NO) and immunoglobulin E (Ig E). Bronchoalveolar lavage fluid (BALF) in each group was collected and the levels of interleukin-10 (IL-10), IL-12 and tumor necrosis factor-α (TNF-α) in the supernatant were measured by ELISA. After centrifugation, the total number of leukocytes and eosinophils (EOS) were counted. The lower right lobe of the lung was cut and ligated for HE staining. Results (1) Cell count: The percentage of total cells and percentage of EOS in BALF were significantly higher in group B than in group A (P <0.05), in group C was significantly lower than in group B (P <0.05) Increased (P <0.05). (2) Compared with group A, the levels of IL-10 and IL-12 in BALF in group B were significantly decreased (P <0.05), and the levels of TNF-α, NO and Ig E in group B were significantly increased ). Compared with group B, the levels of IL-10 and IL-12 in BALF in group C were significantly increased, and the levels of TNF-α, NO and Ig E were significantly decreased. The levels of TNF-α, NO and Ig E in group D were significantly higher (P <0.05). The levels of IL-10 and IL-12 were significantly lower at 1 d, 2 d, 4 d and 7 d. (3) Pathological changes of lung tissue: There was no abnormality in HE staining in group A mice; a large amount of inflammatory cell infiltration was observed in alveolar septum and small vessels in group B; inflammatory cell infiltration in group C was relieved compared with group B; D Part of the mice showed complete occlusion of small airway lumen, inflammatory cell infiltration was significantly worse than the B group. Conclusion rm IL-12 can significantly inhibit the infiltration of inflammatory cells in asthmatic mice and decrease the expression of inflammatory cytokines TNF-α, NO and Ig E, and promote the expression of anti-inflammatory cytokine IL-10, thereby protecting the pathogenesis of asthma effect.