雷帕霉素對哮喘緩解期小鼠模型氣道炎癥的作用

金華良 王利民 王嬌莉 夏俊波 馬勝林

●論 著

雷帕霉素對哮喘緩解期小鼠模型氣道炎癥的作用

金華良 王利民 王嬌莉 夏俊波 馬勝林

目的 探討雷帕霉素對哮喘緩解期小鼠氣道炎癥的作用與機(jī)制。 方法 將 50 只 Balb/c 小鼠隨機(jī)分為 5 組：正常對照組、哮喘緩解組、哮喘模型組、地塞米松組和雷帕霉素組，每組 10 只。采用卵蛋白致敏與激發(fā)制備哮喘模型，休息 3 周后，采用卵蛋白再次激發(fā) 1 周，在小鼠休息期間治療組采用雷帕霉素或地塞米松干預(yù)。采用 Buxco 無創(chuàng)肺功能儀檢測氣道高反應(yīng)性；Luminex 測定肺泡灌洗液 IL-4、IL-5、IL-13、IL-17 及 INF-Y 水平；肺組織 HE 染色評價(jià)觀察小鼠肺組織氣道炎癥；Western blot檢測肺組織 p-S6、S6、AKT、p-AKT（S473）蛋白含量；流式分析測定肺泡灌洗液 CD4+CD25+Foxp3+Treg 細(xì)胞百分率。 結(jié)果 與正常組及哮喘緩解組比較：哮喘模型組氣道反應(yīng)性升高（P＜0.01），灌洗液 IL-4、IL-5、IL-13 及 IL-17 水平增加（均 P＜0.01），肺組織氣道炎癥評分增高（P＜0.01），肺組織 mTORC1 信號通路下游蛋白 p-S6 蛋白增加（P＜0.01），灌洗液 Treg 細(xì)胞比例上升（P＜0.01）；與模型組比較：雷帕霉素在乙酰甲膽堿濃度為 6.25mg/ml時(shí),可降低 Penh 值（P＜0.01），同時(shí)降低肺泡灌洗液 IL-4 水平（P＜0.01），并抑制肺組織炎癥細(xì)胞浸潤（P＜0.01），而肺組織 mTORC1 信號通路下游蛋白 p-S6 蛋白下降（P＜0.01），但雷帕霉素可降低肺泡灌洗液 Treg 細(xì)胞比例（P＜0.01）。 結(jié)論 在哮喘緩解期應(yīng)用雷帕霉素，其可通過 mTORC1 信號通路顯著抑制哮喘急性發(fā)作時(shí)氣道炎癥；在哮喘緩解期應(yīng)用雷帕霉素，對控制哮喘再次發(fā)作具有積極作用。

雷帕霉素 哮喘 mTOR 信號通路

【 Abstract 】 Objective To investigate the effects of rapamycin on airway inflammation in mice with asthma during remission. Methods Fifty Balb/c mice were randomly divided into 5 groups:normal control,asthma,asthma in remission, dexamethasone and rapamycin groups with 10 mice in each group.The asthma model was induced by ovalbumin(OVA) sensitization and challenge;after 3-week interval the mice were challenged by OVA again.Dexamethasone,rapamycin or saline was given to 3 groups,respectively during 3-week interval.Airway reactivity was measured by Buxco's non-invasive system. Cytokines IL-4,IL-5,IL-13,IL-17 and INF-Y in bronchoalveolar lavage fluid(BALF)were assessed by Luminex method.Lung tissues were examined for inflammatory cell infiltration.The expression of p-S6,S6,AKT and p-AKT(S473)in lung tissue was examined by Western blot.CD4+CD25+Foxp3+Treg cells in BALF was assessed by flow cytometry. Results Compared with the normal and remission groups,OVA re-expose significantly increased airway hyperresponsiveness(P＜0.01),elevated IL-4,IL-5, IL-13,IL-17 levels(P＜0.01),and reduced INF-Y in the BALF(P＜0.01);it also markedly increased inflammatory cells in lung tissue,increased Ttreg cells in BALF and decreased expression of p-S6(P ＜0.01).Rapamycin significantly reduced airway hyperresponsiveness to aerosolized methacholine at 6.25mg/ml(P＜0.01),inhibited IL-4 level in BALF,and markedly decreased inflammatory infiltration in lung tissues(P＜0.01).Notably,rapamycin significantly inhibited the expression of p-S6;and also Treg cells in BALF were significantly reduced after rapamycin treatment. Conclusion Antiasthmatic effects of rapamycin during remission time are at least partially through mTORC1 signaling pathway,and rapamycin may be used for asthma patients in remission to control asthma attack.

【 Key words 】 Rapamycin Asthma mTOR signaling pathway

支氣管哮喘是一種以慢性氣道炎癥、氣道高反應(yīng)為特征，伴隨氣道上皮杯狀細(xì)胞增生，黏液高分泌，IgE 顯著增加，以 Th2反應(yīng)為優(yōu)勢的慢性變態(tài)反應(yīng)性疾病[1]。……