Rho/ROCK信号转导通路在哮喘大鼠气道重塑中的作用及干预

The Role of Rho/ROCK Signal Transduction Pathway and the Preventive Effects of Fasudil in the Course of Airway Remodeling of Asthmatic Rats

目的观察法舒地尔对慢性哮喘大鼠镜下气道重塑改变及肺组织RhoA和ROCK I表达的影响,探讨Rho/ROCK信号转导通路在哮喘气道重塑中的作用。方法雄性SD大鼠分为3组,建立慢性哮喘大鼠型,统计支气管肺泡灌洗液(BALF)中细胞计数,观察肺组织病理及超微结构的变化,测定肺内支气管壁厚度(WTt)和支气管平滑肌厚度(WTm),免疫组化法和real-time PCR法分别检测肺组织中RhoA及ROCK I表达情况。结果(1)哮喘组BALF中细胞总数、嗜酸性粒细胞占细胞总数百分比均明显高于正常对照组(均P<0.01);法舒地尔干预后明显抑制炎症细胞浸润,该组BALF中细胞总数较哮喘组不同程度降低(P<0.01)。(2)哮喘组肺组织镜下可见明显炎症性和结构性病理变化,法舒地尔组肺组织炎症反应及重塑改变较哮喘组改善。(3)哮喘组WTt和WTm较正常对照组明显增加(均P<0.01),法舒地尔组WTt和WTm较哮喘组明显降低(均P<0.01)。(4)哮喘组大鼠肺组织RhoA和ROCK I蛋白和mRNA的表达水平明显高于正常对照组(均P<0.01),法舒地尔组明显低于哮喘组(均P<0.01)。(5)WTt、WTm均与大鼠肺组织中ROCK I蛋白水平呈明显正相关(r=0.743、0.974,均P<0.01)。结论哮喘大鼠RhoA和ROCK I表达增高,法舒地尔可能通过影响RhoA和ROCK I的表达,减轻气道炎症及改善气道重塑,表明Rho/ROCK信号转导通路参与哮喘气道重塑的形成。

Objective To study the role of Rho/ROCK signal transduction pathway activating in airway remodeling of rats with asthma by observing the regulation of fasudil to airway remodeling and on the expression of RhoA and ROCK I in the lung tissue. Methods Male rats were randomly divided into three groups. In the experiment, asthma rat model was estab- lished by the ovalbumin （OVA） and AI （OH）3 challenge methods. Leucocyte counting and differential counting in bronchoalveolar lavage fluid （BALF） were performed. The histopathology and ultrastructure changes of pulmonary tissues were observed by light microscope and transmission electron microscope（TEM）. Total bronchial wall thickness（WTt） and airway smooth muscle thickness （WTm） were measured by image-pro plus analysis system. The protein expression of RhoA and ROCK I in rat lung tissue were detected by immunohistochemistry technique（IHC）. The expression of RhoA mRNA and ROCK I mRNA were tested by real-time PCR, Results The total cell numbers in BALF and the ratios of eosinophils to the total cell numbers （EOS%） of the asthma group were all significantly higher than those of normal group（P〈0.01）. The infiltration of inflammatory cells was significantly reduced by using fasudil. The total cell numbers in BALF of the fasudil group were lower than those of asthma group （P〈0.01）. There were conspicuous inflammatory reactions and changes of structure in the lung. Improved histopathological appearance was observed in the lung of fasudil treatment rats. Compared with normal group, WTt and WTm of asthma group significantly increased （all P〈 0.01）. Those of fasudil group were lower than those of asthma group（P〈0.01）. The expression levels of RheA/ROCK I protein and mRNA in lung tissue of the asthma group were significantly elevated compared with those of normal group （all P〈0.01）. Both of them from the fasudil group were significantly lower than those from the asthma group （all P〈0.01）. Linear correlation analysis showed the correlation between WTt and ROCK I protein, WTm and ROCK I protein （r =0.743, 0.974. all P〈0.01）. Conclusion The expression of RhoA and ROCK I in asthmatic rats are increased and fasudil can downregulate the expression of RhoA and ROCK I , relief inflammatory reaction in airway and inhibit the thickening of bronchial wall and smooth muscles, which implicate that Rho/ROCK signal transduction pathway may play an important role in the course of airway remodeling.