孟鲁司特对哮喘小鼠气道炎症及白细胞介素-13和-5的影响

Effects of montelukast on airway inflammation, interleukin-13 and interleukin-5 in asthmatic young mice

目的探讨白三烯受体拮抗剂孟鲁司特通过核转录因子(nuclear transcription factor, NF)-κB-胸腺基质淋巴生成素(thymic stromal lymphopoietin, TSLP)通路对哮喘小鼠气道炎症及对白细胞介素(interleukin, IL)-13、-5的影响。方法 40只4周龄BALB/c雌性小鼠,随机分成正常对照组、模型组和低、中、高剂量组各8只。模型组和低、中、高剂量组采用卵清蛋白+氢氧化铝致敏法诱导哮喘模型。第22天始高、中、低剂量组每次雾化致敏前24 h给予孟鲁司特10、5、2.5mg/kg干预,连续灌喂7d;对照组和模型组于同时间点开始连续7 d给予等量生理盐水。于末次给药后24h取各组肺泡灌洗液进行白细胞分类并计数;采用ELISA法检测各组肺泡灌洗液中IL-13、-5及TSLP水平;取左侧肺组织进行HE、PAS、Masson染色常规病理组织学检查;采用Western blot法检测肺组织细胞质NF-κB p65、IκB及细胞核NF-κB p65的表达水平。结果模型组肺泡灌洗液淋巴细胞、嗜酸性粒细胞、中性粒细胞计数、IL-5、IL-13和TSLP水平高于对照组和低、中、高剂量组(P<0.01),低、中、高剂量组淋巴细胞、嗜酸性粒细胞、中性粒细胞计数和TSLP水平均依次降低(P<0.05);模型组肺组织细胞质NF-κB p65(0.64±0.05)、IκB (0.51±0.04)蛋白表达水平明显低于对照组(1.42±0.09、1.32±0.08)和低剂量组(0.71±0.11、0.62±0.09)、中剂量组(0.81±0.07、0.86±0.12)和高剂量组(0.83±0.09、0.89±0.08)(P<0.05),低、中、高剂量组低于对照组(P<0.05),低剂量组低于中、高剂量组(P<0.05);模型组肺组织细胞核NF-κB p65蛋白(1.32±0.12)表达水平明显高于对照组(0.32±0.03)、中剂量组(0.83±0.11)和高剂量组(0.79±0.08)(P<0.05),低剂量组(1.41±0.13)高于中、高剂量组(P<0.05),低、中、高剂量组高于对照组(P<0.05)。结论孟鲁司特可有效缓解卵清蛋白所致哮喘小鼠的Th2炎性反应,以5mg/kg剂量的效益最好,其作用可能是通过调控NF-κB通路,减少TSLP表达实现的。

Objective To investigate the effects of montelukast on airway inflammation, interleukin(IL)-13 and IL-5 in asthmatic young mice via nuclear transcription factor-κB(NF-κB)-thymic stromal lymphopoietin(TSLP) pathway. Methods Forty female 4-week-old BALB/c mice were randomly divided into normal control group, asthma model group, and montelukast low-, medium-and high-dose groups, with 8 mice in each group. The asthmatic mice models were prepared by ovalbumin + aluminum hydroxide sensitization in model group, and montelukast low-, medium-and high-dose groups. On the 22 nd day, high-, medium-and low-dose groups were given montelukast 10, 5 and 2.5 mg/kg 24 h before atomization sensitization, totally for 7 days. Control group and model group were given the same amount of normal saline. The white blood cells were classified and counted in the alveolar lavage from every group in 24 h after the final administration. The levels of IL-13, IL-5 and TSLP were detected by ELISA technique. The left lung tissue was taken for HE, PAS and Masson staining. The expression of NF-κB p65 and IκB in cytoplasm as well as NF-κB p65 in nucleus were detected by Western blot. Results The lymphocyte, eosinophil and neutrophil counts, as well as the levels of IL-5, IL-13 and TSLP were significantly higher in model group than those in control group, and low-, medium-and high-dose groups(P<0.05), and the lymphocyte, eosinophil and neutrophil counts as well as the TSLP level reduced in low-, medium-and high-dose groups in turn(P<0.05). The levels of NF-κB p65 and IκB in cytoplasm were significantly lower in model group(0.64±0.05, 0.51±0.04) than those in control group(1.42±0.09, 1.32±0.08), low-dose group(0.71±0.11, 0.62±0.09), medium-dose group(0.81±0.07, 0.86±0.12) and high-dose group(0.83±0.09, 0.89±0.08)(P<0.05), lower in low-, medium-and high-dose groups than those in control group(P<0.05), and lower in low-dose group than those in medium-and high-dose groups(P<0.05).The level of NF-κB p65 in nucleus was significantly higher in model group(1.32±0.12)than that in control group(0.32±0.03),medium-dose group(0.83±0.11)and high-dose group(0.79±0.08)(P<0.05),higher in low-dose group(1.41±0.13)than that in medium-and high-dose groups(P<0.05),and higher in low-,medium-and high-dose groups than that in control group(P<0.05).Conclusion Montelukast can effectively alleviate Th2 inflammatory response induced by ovalbumin in young mice with asthma,with the optimal dose of 5 mg/kg,probably by regulating NF-κB pathway and reducing the expression of TSLP.