布地奈德对支气管哮喘小鼠树突细胞胸腺基质淋巴生成素受体表达的影响

The effect of budesonide on thymic stromal lymphopoetin receptor and cytokine profile of dendritic cells in OVA-induced asthma in mice

目的探讨布地奈德对支气管哮喘（简称哮喘）小鼠胸腺基质淋巴生成素（TSLP）-树突细胞（DC）途径和Th2极化的影响及其机制。方法将18只清洁级雌性6～8周龄BALB／c小鼠按随机数字表法分为哮喘组[卵清白蛋白（OVA）腹腔致敏，OVA吸入激发]、布地奈德（BUD）干预组（OVA腹腔致敏，气道内吸入OVA和布地奈德）和对照组（以生理盐水代替OVA混悬液腹腔注射及雾化吸入），每组6只。观察各组小鼠生物学及肺组织病理学改变，收集BALF，并分离培养小鼠脾脏树突细胞，酶联免疫吸附试验（ELISA）检测BALF中TSLP水平、树突细胞上清液中TSLP受体（TSLPR）水平和DC—T细胞共培养上清液中细胞因子。流式细胞仪检测树突细胞表型。多组间比较采用单因素方差分析，组间两两比较采用t检验。结果布地奈德干预组支气管及血管周围炎症细胞浸润轻于哮喘组。哮喘组BALF中TSLP水平[（44．0±5．1）ng／L]和树突细胞培养上清液中TSLPR水平[（19．7±2．2）ng／L]均高于对照组[分别为（14．2±3．6）ng／L和（10．4±1．2）ng／L，t值分别为11．74和9．13，均P〈0．01]，BALF中TSLP水平与IL4水平和嗜酸粒细胞呈正相关（r值分别为0．89和0．82，均P〈0．05）；布地奈德干预组BALF中TSLP水平[（19．2±5．8）ng／L]及树突细胞表达TSLPR水平[（12．5±2．8）ng／L]均低于哮喘组[分别为（44．0±5．1）ng／L和（19．7±2．2）ng／L]，差异有统计学意义（t值分别为-7．86和-4．97，均P〈0．01）；布地奈德干预组BALF及DC—T细胞共培养上清液中IL-5[分别为（41±4）ng／L和（28±9）ng／L]低于哮喘组[分别为（81±8）ng／L和（51±14）ng／L]，差异有统计学意义（t值分别为-11．00和-3．35，均P〈0．01），3组间IFN．叮差异无统计学意义（F值分别为0．97和0．82，均P〉0．05）；布地奈德干预组与哮喘组相比CD。、CD。和CD。等树突细胞表型下降。结论布地奈德影响树突细胞表型，下调BALF中TSLP水平和树突细胞表达TSLPR水平，可能通过影响TSLP—DC途径而抑制哮喘Th2极化反应。

Objective To investigate the effect of budesonide （BUD） on thymic stromal lymphopoetin receptor （TSLPR） of dendritic cells （ DCs ） in OVA-induced mouse asthma models and to explore the mechanisms by studying the effect of BUD on function of DCs from the model. Methods Eighteen BALB/e female mice were randomly divided into a control group, an asthma group and a BUD intervention group, with 6 mice in each group. Mice were sensitized and challenged with ovalhumin （OVA） to establish the asthmatic model. The bronchoalveolar lavage fluid （BALF） and DCs of spleen from the 3 groups were harvested and the supernatants of BALF and DCs were analyzed for levels of TSLP and TSLPR, respectively, by commercially available ELISA kit. The percentage of eosinophils （EOS） in BALF was counted. The expression of CD40, CDs0 and CD86 in DCs was detected by FACS. The DCs were then washed, and co-cultured in vitro with autologous T cells purified by a nylon cotton column. The supernatants of DC-T co-culture were collected after 72 h incubation, and analyzed for levels of interleukin-5 （lL-5） and interferon-γ （IFN-γ） by ELISA. Results The levels of TSLP in BALF and TSLPR in DCs from the asthma group were significantly increased compared with the control group [ （44. 0 ± 5.1 ） ng/L vs （14. 2 ± 3.6） rig/L, P 〈 0. 01 and （ 19.7 ± 2.2） ng/L vs （ 10.4 ± 1.2） rig/L, P 〈 0.05, respectively ]. The expression of CD40, CD80 and CD86 of DEs and IL-5 in the culture supernatants of DC-T co-culture was significant up- regulated in the asthma group compared with the control group （ P 〈 0. 05 ）. Furthermore, the addition of BUD reduced the expression of CD40, CD80, CD86 , TSLPR in DCs, IL-5 in the culture supernatants of DC- T co-culture, TSLP and EOS in BALF. The level of INF-γ in the DC-T co-culture supernatants of the 3 groups did not achieve statistical significance （ F = 0. 82, P 〉 0. 05 ） . Conclusion These results demonstrate that the therapeutic activity of BUD in asthmatic mice may be related to modulation of Thl and Th2 cell functions and this effect is probably mediated through the TSLP-DC pathway.