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目的:观察黄芪多糖对大鼠小肠上皮细胞(IEC-6)迁移多胺信号通路的影响,探讨益气健脾中药黄芪促进胃肠黏膜损伤修复的作用机制。方法:在划痕损伤致细胞迁移模型上观察黄芪多糖对细胞迁移的影响;流式细胞仪检测细胞膜电位(Em)和细胞内游离钙离子水平([Ca2+]cyt);Western blot法检测钾通道蛋白Kv1.1和RhoGTPaes(Cdc42、Rac1、Rho A)蛋白表达。结果:黄芪多糖(20、40、80mg/L,下同)有以下作用:(1)促进细胞迁移(P<0.01),逆转α-二氟甲基鸟氨酸(DFMO)所致的细胞迁移抑制(P<0.01);(2)提高细胞迁移过程Kv1.1蛋白表达、逆转DFMO所致的Kv1.1蛋白表达抑制(P<0.05,P<0.01);(3)促进Em超极化、逆转DFMO所致的Em去极化(P<0.05,P<0.01);(4)提高细胞迁移过程[Ca2+]cyt、逆转DFMO所致的[Ca2+]cyt降低(P<0.01);(5)提高细胞迁移过程Cdc42、Rac1、Rho A蛋白表达、逆转DFMO的抑制作用(P<0.05,P<0.01)。结论:黄芪多糖促进IEC-6细胞迁移与影响多胺信号通路相关指标有关。
Objective: To observe the effect of Astragalus Polysaccharides on the migration of polyamine signal pathway in rat intestinal epithelial cells (IEC-6), and to explore the mechanism of Astragalus membranaceus in promoting the repair of gastrointestinal mucosal injury. Methods: The effect of APS on cell migration was observed on scratch-induced cell migration model. Cell membrane potential (Em) and intracellular free calcium level ([Ca2 +] cyt) were detected by flow cytometry. Protein Kv1.1 and RhoGTPaes (Cdc42, Rac1, Rho A) protein expression. Results: Astragalus polysaccharides (20, 40, 80mg / L, the same below) had the following effects: (1) promoted cell migration (P <0.01) and reversed cell migration induced by α-difluoromethylornithine (DFMO) (P <0.01). (2) The expression of Kv1.1 protein in cell migration was increased, and the inhibition of Kv1.1 protein expression induced by DFMO was reversed (P <0.05, P <0.01) Reverse DFMO induced depolarization of Em (P <0.05, P <0.01); (4) increased [Ca2 +] cyt cell migration and reduced [Ca2 +] cyt caused by DFMO (P < Increase the protein expression of Cdc42, Rac1 and Rho A, and reverse the inhibitory effect of DFMO (P <0.05, P <0.01). Conclusion: Astragalus polysaccharide can promote the migration of IEC-6 cells and the related indicators of polyamine signaling pathway.