基于MAL调控CADM1的转运探究柴朴汤对肝郁型哮喘小鼠气道上皮细胞的影响及分子机制

Applicants found that the effective rate of Chaipu Decocton in relieving acute asthma attack was 93.1% through preliminary clinical experiments, but its mechanism action was unknown. In preliminary experiments, it was found that Chaipu Decocton regulate the expression of T-lymphocyte maturation associated protein (MAL), and MAL may participate the development of asthma through its vesicle transport function. Therefore, the following scientific hypothesis is proposed: Chaipu Decocton can upregulate the expression of MAL, maintain the normal morphology and function of epithelial cells, regulate the transportation and secretion of the cell membrane-promoting cell adhesion molecule (CADM1), and delay the inflammatory response of liver-depression asthma. This project intends to use MAL knockout gene asthma mice and epithelial cell lines as models for treatment, study the effect of Chaipu Decocton on the morphology and function of epithelial cells at the anima level and study the effect and molecular mechanism of Chaipu Decocton containing serum on airway epithelial MAL from the cell level. The effect of Chaipu Decocton-containing serum on the airway epithelial MALs was studied from the cellular level, and the mechanism was analyzed to find whether the Chaipu Decocton's mechanism action on asthma was related to the transport and secretion of MAL-regulated apical membrane proteins. The completion of the project can further clarify the role of Chaipu Decocton in the treatment of liver-depression asthma and related molecular mechanisms.

申请者通过前期临床实验发现，柴朴汤缓解哮喘急性发作的有效率达到93.1%，但其机制尚不明确。在预实验中发现，柴朴汤能调控T-淋巴细胞相关蛋白（T-lymphocyte maturation associated protein ,MAL）的表达，并且MAL可能通过其囊泡转运功能参与哮喘的发展过程。因此提出以下科学假说：柴朴汤可上调MAL的表达，维持上皮细胞的正常形态和功能，调控顶端膜蛋白细胞粘附分子CADM1的转运和分泌，延缓肝郁型哮喘的炎症反应。本项目拟以MAL敲基因肝郁型哮喘小鼠和上皮细胞系为模型，采用柴朴汤进行处理,分别在整体动物水平研究柴朴汤对上皮细胞形态和功能的影响，从细胞水平探讨柴朴汤含药血清对气道上皮MAL的影响和分子机制，进而解析柴朴汤对肝郁型哮喘的作用机制是否与MAL调控顶端膜蛋白转运和分泌相关。该项目的完成可以进一步阐明柴朴汤治疗哮喘的作用及相关分子机制。

气道上皮细胞是机体抵抗外源病原微生物侵袭的首要屏障，细胞结构和功能的完整性的维持是生命状态安全的保障，上皮细胞脱落损伤是呼吸道疾病如哮喘发生发展的关键潜在机制。MAL（T细胞分化晚期蛋白）包含四个潜在的跨膜结构域，类似于许多膜蛋白的结构，证实可控制合成蛋白从高尔基体向上皮细胞顶端的极化运输，与维持细胞的极性息息相关。通过对MAL基因细化研究发现，哮喘和肝郁型哮喘模型小鼠，MAL基因表达量呈明显下降趋势，且肝郁型下降更明显。CADM1（细胞粘附分子1）作为细胞间粘附分子，表达也同时下调，但两者无明显相关性。柴朴汤方剂干预，MAL和CADM1表达上升。进一步建立MAL基因敲除小鼠模型，哮喘和抑郁症造膜后，与野生组相比，CADM1和炎症因子表达量下降，柴朴汤干预治疗后结果无明显差异，提示MAL基因是柴朴汤治疗哮喘的生物学靶点之一。