五苓散同方汤剂缺失成分对HK-2细胞AQPs表达的影响

Research on the Chemical Components Deficiency and the Corresponding Efficacy Missing in the Decoction of Wuling San Comparing with the Powder Form

目的研究五苓散同方汤剂缺失成分对人肾小管上皮细胞(Human renal tubular epithelial cell line, HK-2)AQP1和AQP4表达的调节作用,进而探讨五苓方宜散不宜汤的作用机制。方法采用MTT法检测麦角甾醇(ergosterol, ERG)、香豆素(coumarin, COU)及23-乙酰泽泻醇B(alisol b 23-acetate,ALI)三种汤剂缺失成分对HK-2细胞的抑制率情况,乳酸脱氢酶漏出率(LDH)反映ERG、COU及ALI对HK-2细胞的损伤程度,RT-PCR及Western Blot法测定ERG、COU及ALI对HK-2细胞AQP1和AQP4表达的影响。结果 ERG、COU及ALI均能显著降低HK-2细胞AQP1和AQP4 mRNA及蛋白的表达。结论五苓方宜散不宜汤的作用机制与其汤剂药效成分损失密切相关。

Objective To investigate the effect of three missing components on AQP1 and AQP4 expression regulation in Human renal tubular epithelial cells,then to explore the mechanism of the powder of Wuling San better than the decoction form.Methods The inhibition of cell proliferation is examined by MTT assay,the Cytotoxicity is examined by LDH leakage rate,and the expression of AQP1 and AQP4 proteins are determined by Western blot,as the same time,the expression of AQP1 and AQP4 mRNA is determined by Real-time PCR.Results Three compounds of ergosterol,coumarin and alisol b 23-acetate are extracted from the powder of Wuling San,but lost in decoction of Wuling San which significantly decreased levels of AQP1 and AQP4 mRNA and protein in HK-2 cells.Conclusion The powder of Wuling San exerts better renal water regulation effect by modulating levels of AQP1 and AQP4 expression in HK-2 cell than its decoctive form.The molecular mechanism of its action may be associated with the loss of a large number of chemical compounds in the decoction of Wuling San.