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目的观察白术提取物对小肠上皮细胞(IEC-6)迁移多胺介导钾通道激活信号通路指标Rho GTPaes(Rho A、Rac1和Cdc42)表达的影响,对肌球蛋白II(myosinⅡ)分布和表达的影响,以探讨益气健脾中药白术对胃肠黏膜损伤修复的作用机制。方法荧光定量PCR法检测Rho A、Rac1和Cdc42 mRNA表达;Western blot法检测Rho A、Rac1和Cdc42蛋白表达;免疫荧光法检测myosinⅡ蛋白分布和表达。结果白术提取物能提高细胞迁移过程Rho A、Rac1、Cdc42 mRNA和蛋白表达,逆转多胺合成抑制剂二氟甲基鸟氨酸(DFMO)所致的Rho A、Rac1、Cdc42 mRNA和蛋白表达抑制;提高细胞迁移过程myosinⅡ表达,从而增加细胞骨架应力纤维形成,改善DFMO所致的myosinⅡ蛋白表达和分布的抑制。结论白术提取物促进IEC-6细胞迁移的作用机制,与其影响多胺介导钾通道激活信号通路有关。
Objective To observe the effects of Atractylodes macrocephalae extract on the expression of Rho GTPaes (Rho A, Rac1 and Cdc42), which is the marker of potassium channel activation induced by migration of small intestine epithelial cells (IEC-6), the distribution and expression of myosinⅡ In order to investigate the mechanism of action of Atractylodes Rhizoma Atractylodis Macrocephalae on the repair of gastrointestinal mucosal injury. Methods The mRNA expressions of Rho A, Rac1 and Cdc42 were detected by real-time PCR. The expressions of Rho A, Rac1 and Cdc42 were detected by Western blot. The distribution and expression of myosin II protein were detected by immunofluorescence. Results The extract of Atractylodes macrocephala increased mRNA and protein expression of Rho A, Rac1 and Cdc42 during cell migration and reversed the mRNA and protein expression of Rho A, Rac1 and Cdc42 induced by polyamine synthesis inhibitor difluoromethylornithine (DFMO) ; Increased myosin Ⅱ expression during cell migration, increased the formation of cytoskeletal stress fibers, and inhibited the DFMO-induced myosinⅡ protein expression and distribution. Conclusion Atractylodes macrocephalae extract can promote the migration of IEC-6 cells, which may be related to the effect of polyamine on the activation of potassium channel.