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为了研究沉默c-myc基因后Raji细胞自噬活性的变化及其机制,采用脂质体转染SiRNA法沉默c-myc基因,在透射电镜下观察细胞自噬体的形成情况,采用western blot法测定LC3-Ⅰ、LC3-Ⅱ、Beclin-1、磷酸化P70、磷酸化AKT蛋白的表达。研究发现沉默c-myc基因后,电镜下观察到Raji细胞自噬活性随时间而增强,LC3-Ⅰ、LC3-Ⅱ蛋白表达、LC3-Ⅱ/LC3-Ⅰ比值随时间而增高;Beclin-1表达增高,磷酸化P70表达降低,磷酸化AKT表达无明显变化。结果提示沉默c-myc基因使Raji细胞自噬活性明显增强,其机制是通过上调Beclin-1表达和抑制mTOR的活性来实现的。
In order to study the changes of autophagic activity of Raji cells after silencing c-myc gene and its mechanism, the c-myc gene was silenced by liposome transfection with SiRNA, and the formation of autophagosomes was observed under transmission electron microscope. Western blot The expression of LC3-I, LC3-II, Beclin-1, phosphorylated P70 and phosphorylated AKT protein was determined. After silencing c-myc gene, the autophagy activity of Raji cells was observed under electron microscope. The expression of LC3-Ⅰ, LC3-Ⅱ and the ratio of LC3-Ⅱ / LC3-Ⅰ increased with the time; the expression of Beclin-1 Increased phosphorylated P70 expression decreased, phosphorylated AKT expression no significant change. The results suggest that silencing of c-myc gene enhances the autophagy activity of Raji cells by up-regulating the expression of Beclin-1 and inhibiting the activity of mTOR.