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目的:观察青钱柳多糖对肝细胞胰岛素信号传导通路关键靶点基因、蛋白表达的影响。方法:以H4IIE大鼠肝细胞为模型,培养72 h后,采用Neutral Red法检测青钱柳多糖(50、100、200、400μg·m L-1)对细胞活性的影响;根据细胞活性检测结果分为正常对照组(Control组)、胰岛素组(Insulin组)、青钱柳多糖低剂量组(LCP组)和高剂量组(HCP组);药物干预2 h后RT-PCR检测肝细胞相应基因表达,药物干预30 min后,利用Western blot法检测肝细胞相应蛋白磷酸化水平。结果:与Control组比较,100、200、400μg·m L-1青钱柳多糖干预后肝细胞活性显著升高(P<0.01);干预2 h后,LCP组Ins R、IRS-2基因表达显著升高(P<0.05),Insulin组和HCP组Ins R、IRS-2基因表达极显著升高(P<0.01);干预30 min后,Insulin组、HCP组Ins Rβ、IRS-2、Akt蛋白磷酸化水平显著升高(P<0.01)。结论:青钱柳多糖具有增加肝细胞活性作用,其上调肝细胞胰岛素信号通路关键靶点Ins R、IRS-2基因表达,以及Ins Rβ、IRS-2、Akt蛋白磷酸化水平可能是其发挥降糖作用的机制。
Objective: To observe the effects of Cyclocarya paliurus polysaccharides on the expression of genes and proteins of key hepatic insulin signaling pathways. Methods: H4IIE rat hepatocytes were used as a model. After cultured for 72 h, the effects of Cyclocarya paliurus polysaccharides (50, 100, 200, 400 μg · m L-1) on cell viability were determined by Neutral Red assay. The rats in Control group, Insulin group, LCP group and HCP group were divided into two groups: control group, Hp group, Insulin group, Expression and drug intervention for 30 min, Western blot was used to detect the corresponding protein phosphorylation of hepatocytes. Results: Compared with Control group, the activities of hepatocytes in 100, 200, 400μg · m L-1 Cyclocarya paliurus polysaccharide increased significantly (P <0.01). After 2 h intervention, the expression of Ins R, IRS- (P <0.05). The Ins IR and IRS-2 gene expressions in Insulin group and HCP group were significantly higher than those in Insulin group and HCP group (P <0.01) Protein phosphorylation was significantly increased (P <0.01). CONCLUSION: Cyclocarya paliurus polysaccharides can increase hepatic cell viability and increase the expression of insulin receptor IRS-2 and Ins Rβ, IRS-2, Akt protein phosphorylation at the key targets of hepatocyte insulin signaling pathway. Mechanism of sugar action.