TSLP在哮喘小鼠气道黏液高分泌中的作用

发布时间：2018-06-03 12:37

  本文选题：哮喘 + 动物模型　； 参考：《遵义医学院》2012年硕士论文

【摘要】：目的：评价以鸡卵清白蛋白(OVA)致敏和激发小鼠制备哮喘模型。 方法：清洁级BALB/c小鼠20只随机分为哮喘组(AS组)和生理盐水对照组(NS组),每组10只。AS组小鼠于第1、13d以OVA致敏,从第1次致敏后第19d开始以5%OVA溶液雾化激发并连续5d,末次激发24h后处死小鼠；NS组以生理盐水代替OVA作对照,余处理同AS组。检测两组小鼠支气管肺泡灌洗液(BALF)中细胞总数和细胞分类计数,采用酶联免疫吸附试验(ELISA)检测BALF白细胞介素5(IL-5)和IL-32水平的变化,苏木素-伊红(HE)染色观察肺组织病理学改变,阿辛蓝-过碘酸雪夫(AB-PAS)染色观察气道上皮杯状细胞及黏液物质改变,Van Gieson(VG)染色观察气道周围胶原纤维增生情况,免疫组织化学(IHC)检测肺组织中黏蛋白5ac(Muc5ac)和IL-32蛋白的表达水平,实时荧光定量逆转录PCR和免疫印迹法(Western blot)分别检测肺组织中Muc5ac mRNA和蛋白的表达水平。 结果：AS组小鼠肺组织中出现哮喘特征性病理改变,BALF中细胞总数、嗜酸性粒细胞、淋巴细胞、中性粒细胞百分比、IL-5和IL-32水平、气道炎症病理评分、气道上皮杯状细胞及黏液物质阳性相对着色面积、气道周围胶原纤维阳性相对着色面积、肺组织IL-32蛋白、Muc5ac蛋白和Muc5ac mRNA表达均高于NS组,差异有统计学意义(P0.05)。 结论：以OVA致敏和激发小鼠可以成功复制哮喘小鼠动物模型；哮喘小鼠出现明显的气道炎症、气道上皮杯状细胞增生／化生及气道黏液高分泌；哮喘小鼠气道炎症可能导致气道黏液高分泌的发生 目的：通过应用罗格列酮和地塞米松干预哮喘小鼠,探讨胸腺基质淋巴细胞生成素(TSLP)在哮喘小鼠气道黏液高分泌中的作用。 方法：清洁级BALB/c小鼠40只随机分成生理盐水对照组(NS组)、哮喘组(AS组)、罗格列酮干预组(AS+ROS组)、地塞米松干预组(AS+DEX组),各10只。AS组和NS组处理同第一部分；AS+ROS组从第1次致敏后第17d开始予罗格列酮5mg*kg-1*d-1灌胃,第19d开始,于每日雾化激发前30min给药,连续7d,余处理同AS组；AS+DEX组于每日雾化激发前30min予地塞米松2mg/kg腹腔注射,连续5d,余处理同AS组。测定各组小鼠BALF中细胞总数和细胞分类计数,采用ELISA检测BALF中IL-5水平的变化,HE染色观察肺组织病理学改变,AB-PAS染色观察气道上皮杯状细胞及黏液物质改变,VG染色观察气道周围胶原纤维增生情况,IHC检测肺组织中Muc5ac、TSLP、CD11。和脾组织中CD11。蛋白的表达水平,实时荧光定量逆转录PCR和Western blot分别检测肺组织中Muc5ac、TSLP mRNA和蛋白的表达水平。 结果：AS组BALF细胞总数、嗜酸性粒细胞、淋巴细胞、中性粒细胞百分比、IL-5水平、气道炎症病理评分、气道上皮杯状细胞及黏液物质阳性相对着色面积、气道周围胶原纤维阳性相对着色面积、肺组织和脾组织中CD11C蛋白、肺组织中Muc5ac、 TSLP蛋白及Muc5ac、TSLP mRNA表达均较NS组、AS+ROS组、AS+DEX组增高,差异有统计学意义(P0.05)。 结论：哮喘小鼠气道上皮细胞中TSLP表达增高,TSLP可能通过激活CD11c+DCs诱导CD4+T细胞前体向Th2细胞分化,启动Th2细胞优势免疫应答,加重哮喘气道炎症,导致气道Muc5ac mRNA和蛋白的高表达,促进气道黏液高分泌；罗格列酮和地塞米松可能通过抑制肺组织中TSLP勺表达,减少Th2细胞因子的分泌,从而减轻哮喘小鼠的气道炎症和气道黏液高分泌。
[Abstract]:Objective : To evaluate the effect of OVA on the preparation of asthma model .

Methods : 20 BALB / c mice were randomly divided into two groups : asthma group ( AS group ) and normal saline control group ( NS group ) . Each group of 10 . AS group mice were sensitized with OVA on Days 1 and 13d . After the first sensitization , the mice were killed at 5 % OVA solution for 5 days , and the mice were sacrificed at the last 24 hours ;
The expression levels of Muc5ac ( Muc5ac ) and IL - 32 in lung tissues were observed by enzyme - linked immunosorbent assay ( ELISA ) , and the expression level of Muc5ac ( Muc5ac ) and IL - 32 protein in lung tissues were detected by immunohistochemistry ( IHC ) , and the level of Muc5ac mRNA and protein expression in lung tissues was detected by real - time fluorescence quantitative reverse transcription - PCR and Western blot .

Results : The expression of IL - 32 , Muc5ac and Muc5ac mRNA in BALF were higher than that in NS group ( P0.05 ) .

Conclusion : OVA - sensitized and challenged mice can successfully replicate the mouse model of asthma mice .
In asthmatic mice , airway inflammation , airway epithelial cup - like cell proliferation / chemical and airway mucus hypersecretion were observed .
Airway inflammation in asthmatic mice may cause airway mucus hypersecretion

Objective : To study the role of thymic stromal lymphopoiesis ( TSLP ) in the airway mucus hypersecretion of asthmatic mice by the intervention of roglione and dexamethasone in the treatment of asthma mice .

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