黄芪多糖通过AQP5调节ERK/MLCK通路干预肠黏膜通透性改善脾虚水湿运化失常机制研究

脾虚水湿运化失常可引发各种病证或出现在疾病进展的不同阶段，并使其迁延不愈。脾虚失运是脾虚水湿运化失常的基本病机特征，与肠黏膜改变关系密切。我们前期研究表明，黄芪多糖可明显改善脾虚水湿不化大鼠肠道功能及水液代谢障碍，上调AQP5低表达。AQP5既促进水液排泌又可诱导ERK/MLCK通路增强紧密连接降低肠黏膜通透性。而据此提出科学假说：“黄芪多糖通过AQP5诱导的ERK/MLCK通路增强紧密连接，降低肠黏膜通透性，改善脾虚水湿运化失常”。本研究拟使用脾虚水湿不化动物模型和AQP5沉默细胞模型，采用基因转染、qRT-PCR、蛋白印迹等方法，研究黄芪多糖对AQP5低表达诱导ERK/MLCK通路激活破坏紧密连接增加肠黏膜通透性而导致脾虚水湿运化失常的调节作用，并使用ERK抑制剂进行反证，为揭示黄芪多糖改善脾虚水湿运化失常的分子机制提供实验依据和研究思路。

Dysfunction of dampness transportation due to spleen deficiency can lead to various diseases or occur at different stages of disease progression and make its recurrent and persistent. Spleen deficiency is the basic pathogenesis of dysfunction of dampness transportation, which related to intestinal mucosa changes. Our previous studies have shown that Astragalus polysaccharides can significantly improve disorders of the intestinal function and water metabolism in rats with dampness stagnancy due to spleen deficiency syndrome, up-regulate low expression of AQP5. AQP5 not only promotes water and fluid excretion, but also induces ERK/MLCK pathway to enhance tight junction and decrease intestinal mucosal permeability. Therefore, we propose the scientific hypothesis: "Astragalus polysaccharide enhances tight junction and reduces intestinal mucosal permeability to attenuate dysfunction of dampness transportation due to spleen deficiency via ERK/MLCK pathway mediated by AQP5". In this study, the rats model with dampness stagnancy due to spleen deficiency syndrome and AQP5 silencing cells are established, gene transfection, qRT-PCR, western blot will be applied to study the regulatory effect of Astragalus polysaccharide on dysfunction of dampness transportation due to spleen deficiency caused by tight junction reduction and intestinal permeability increase because of AQP5-induced activation of ERK/MLCK pathway, and ERK inhibitor will be applied to verified that. This study will provide new research idea for revealing the mechanism action of Astragalus Polysaccharide in attenuating dysfunction of dampness transportation due to spleen deficiency.