从水通道蛋白1介导的细胞迁移探讨“肾主水”的分子机制

TCM zangxiang Theory deams that kidney governs water, that is, kidney qi is in charge of regulating the whole body water metabolism. Medical research suggests the function and regulatory mechanism of aquaporin (AQP) have close relation with water metabolism. In the aquaporin family, AQP1 under the influence of corticosteroid plays important role in urine concentration, mediates cell migration for the recovery of renal injury. Our preliminary studies confirmed that the kidney warming prescriptions can enhance the function of the HPA axis in kidney yang deficient rats, promote the secretion of glucocorticoid and increase the expression of AQP1. It is estimated that the expression of AQP1 in kidney may be regulated by corticosteroid through HPA axis, which thereby influences the migration of renal cells, facilitates the repair of injured kidney tissue and contributes to the formation of urine. All of this process may biologically underlie the mechanism of "kidney governs water". Out program aims to observe the alteration of HPA axis in kidney yang deficient rats, to detect the expression of AQP1 and migration in proximal tubular epithelial cells and straight vascular cells, and therefore confirm the regulatory process of cell migration and renal repair with the kidney warming prescriptions under the guidance of TCM zangxiang Theory. It is hopeful to provide the basic evidence for clarification of scientific connotation of "kidney governs water", and at the time provide a reference for mechanism investigation in the treatment of kidney yang deficiency and metabolic disorders with the kidney warming prescriptions.

中医藏象理论认为，"肾主水"，即肾气主司和调节全身水液代谢。现代医学研究表明，水通道蛋白(AQP)的功能及其调节机制与机体水液代谢密切相关。其中，肾脏AQP1不仅在尿浓缩机制中起重要作用，且其介导的细胞迁移关乎肾脏损伤修复，而其表达则受皮质激素的调控。我们前期研究证实，温肾药能增强肾阳虚大鼠HPA轴的功能，促进糖皮质激素分泌与AQP1表达。由此推测，HPA轴通过皮质激素调控肾脏AQP1的表达，继而调节肾脏细胞的迁移能力，促进肾组织损伤修复与尿液生成功能，这可能是"肾主水"的重要生物学基础。本项目拟在藏象理论指导下，观察肾阳虚模型大鼠HPA轴的变化，肾脏近端小管上皮细胞与直小血管细胞的AQP1表达、细胞迁移及肾脏损伤修复状况，并观察温肾药肾气丸的干预作用，"以方测证"。为初步阐明"肾主水"理论的科学内涵提供依据，同时为研究温肾方药治疗肾阳虚证及其水液代谢紊乱的作用机制提供参考。

中医藏象理论认为，“肾主水”，即肾气主司和调节全身水液代谢。现代医学研究表明，水通道蛋白(AQP)的功能及其调节机制与机体水液代谢密切相关。其中肾脏 AQP1不仅在尿浓缩机制中起重要作用，且其介导的细胞迁移关乎肾脏损伤修复，而其表达则受皮质激素的调控。因此本项目拟在藏象理论指导下，观察肾阳虚模型大鼠 HPA 轴的变化，肾小管上皮细胞AQP1 的表达、细胞迁移及肾脏损伤修复状况，并用温肾药肾气丸“以方测证”。为初步阐明“肾主水”理论的科学内涵提供实验依据。. 主要研究内容：①采用不同剂量腺嘌呤建立肾阳虚证肾损伤大鼠模型，观察大鼠肾阳虚证一般症状体征、肾功能及肾阳虚相关指标，明确不同剂量腺嘌呤造模的特点，筛选了肾损伤程度适当的肾阳虚证模型的造模剂量。②观察了肾阳虚大鼠 HPA 轴的功能、糖皮质激素分泌、组织病理变化以及肾脏 AQP1 的表达变化及肾气丸的干预作用。③体外培养肾小管上皮细胞，观察AQP1的表达与细胞迁移效应及肾气丸的干预作用。④结合整体动物实验和体外细胞实验，观察了肾气丸抗肾阳虚证大鼠肾间质纤维化的作用，初步探讨其作用机制。. 研究结果显示：①腺嘌呤150mg/kg持续给予14d所制备的肾阳虚模型，肾脏损伤程度适中，肾气丸干预后模型各指标改善较明显，是适合进一步研究“肾主水”的肾阳虚证模型。②肾气丸能显著改善腺嘌呤所致肾阳虚大鼠的肾功能、HPA轴相关指标和肾小管周毛细血管的损伤，提高AQP1 mRNA水平和蛋白表达。③肾气丸含药血清能抑制AQP1 RNAi慢病毒感染NRK-52E细胞后引起的AQP1 mRNA水平和蛋白表达下降以及延缓NRK-52E细胞粘附、迁移和促进损伤愈合。④肾气丸可能通过调控由TGF-β1介导的肾小管上皮细胞EMT信号通路中的Vimentin、p-Smad2/3、p-GSK-3β等多个位点，抑制EMT过程从而延缓肾间质纤维化进程。. 以上研究表明，温肾药肾气丸可能是通过HPA轴调控AQP1的表达、促进细胞迁移而修复肾损伤，从而改善肾阳虚证水液代谢紊乱，初步揭示了“HPA轴-肾脏AQP1-细胞迁移-肾损伤修复”为“肾主水”的分子机制之一。